Methods and compositions for treating infection

ABSTRACT

Provided herein are compositions and methods for treating or preventing infection.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application No.61/706,492, filed on Sep. 27, 2012, which is incorporated by referenceherein in its entirety.

STATEMENT REGARDING FEDERALLY FUNDED RESEARCH

This invention was made with government funding under Grant Nos.1R01AI1075033-03 and 5P50GM069663 from the National Institutes ofHealth. The government has certain rights in this invention.

BACKGROUND

Infectious diseases affect the health of people and animals around theworld, causing serious illness and death. Thus, an urgent need existsfor treatments for infections.

SUMMARY

Provided herein is a method of treating or preventing an infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject

a compound of Formula I

wherein R represents hydrogen or hydroxyl and R¹ represents hydrogen, orR and R¹ taken together form a second bond between the carbon atomsbearing R and R¹; R² represents hydrogen or phenyl; n is zero or apositive whole integer of from 1 to 4; X represents CH₂, CHOH, NH, C═O,CHNR³R⁴, where R³ and R⁴ independently are hydrogen or lower alkyl; andZ represents thienyl, pyridinyl, substituted pyridinyl, phenyl orsubstituted phenyl wherein the substituents on the substituted pyridinylor substituted phenyl are selected from phenyl, pyridinyl, nitro, ahalogen atom, such as chlorine, fluorine, bromine, or iodine, a straightor branched lower alkyl chain of from 1 to 4 carbon atoms, a loweralkoxy group of from 1 to 4 carbon atoms, amino, a mono ordi(lower)alkylamino group, a saturated monocyclic heterocyclic groupsuch as pyrrolidino, piperidino, morpholino, orN-(lower)alkylpiperazino, or a group having the structure —COOR⁵,—CR⁶R⁷COOR⁵, —CF₃, CHF₂, CH₂F, or

where R⁵ is hydrogen or lower alkyl, and R⁶ and R⁷ independently arehydrogen or methyl or a pharmaceutically acceptable salt thereof.

Further provided is a method of treating or preventing an infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject

a compound of Formula III

or a pharmaceutically acceptable salt thereof, wherein R¹ and R² areindependently selected from ethyl or methyl, n 1 or 2, R³ and R⁴ areboth phenyl or substituted phenyl, wherein the substituent can be halo(for example, fluoro-, chloro-, iodo- or bromo-), hydroxyl, a loweralkyl or a substituted lower alkyl wherein the substituent can be halo,hydroxy, or lower alkoxy, and R⁵ is hydrogen, a halogen, a lower alkylfrom about 1 to 4 carbon atoms or a substituted alkyl wherein thesubstituent can be halo, hydroxy, or lower alkoxy.

Also provided is a method of treating or preventing infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of:

-   -   a compound of Formula V

wherein R₁ is a lower alkyl group having from 1 to 4 carbon atoms beingsubstituted with one or several halogen atoms, Z is hydrogen or a loweralkyl group having from 1 to 4 carbon atoms, m is 2 or 3 and X₂ is theethylene imino group or the group having the formula:

wherein R is hydrogen or a lower alkyl group having from 1 to 4 carbonatoms which can be substituted with a chlorine atom or a hydroxy group,and Z and m have the above-given meaning,

-   -   a compound of Formula X

wherein R represents a lower alkyl or a lower alkenyl group, a phenylgroup which can be substituted by halogen, methyl or lower alkoxygroups, or a benzyl group which can be nuclear substituted by halogen,methyl or lower alkoxy groups,X represents —O—, —S—, —SO—, or —SO₂—,n represents an integer from 1-4, and

Aryl represents a phenyl group which can be substituted by lower alkoxyor lower alkylmercapto groups;

-   -   a compound of Formula XII

wherein R and R⁶ are the same or different and are hydroxy, loweralkoxy, lower alkenoxy, dilower alkylamino lower alkoxy(dimethylaminoethoxy), acylamino lower alkoxy (acetylaminoethoxy),acyloxy lower alkoxy (pivaloyloxymethoxy), aryloxy, such as phenoxy,arloweralkoxy, such as benzyloxy, substituted aryloxy or substitutedarloweralkoxy wherein the substitutent is methyl, halo or methoxy,amino, loweralkylamino, diloweralkylamino, hydroxyamino,arloweralkylamino such as benzylamino;

R¹ is hydrogen, alkyl of from 1 to 20 carbon atoms which includebranched and cyclic and unsaturated (such as allyl) alkyl groups,substituted loweralkyl wherein the substituent can be halo, hydroxy,lower alkoxy, aryloxy such as phenoxy, amino, diloweralkylamino,acylamino, such as acetamido and benzamido, arylamino, guanidino,imidazolyl, indolyl, mercapto, loweralkylthio, arylthio such asphenylthio, carboxy or carboxamido, carboloweralkoxy, aryl such asphenyl or naphthyl, substituted aryl such as phenyl wherein thesubstituent is lower alkyl, lower alkoxy or halo, arloweralkyl,arloweralkenyl, heteroarlower alkyl or heteroarlower alkenyl such asbenzyl, styryl or indolyl ethyl, substituted arloweralkyl, substitutedarloweralkenyl, substituted heteroarlower alkyl, or substitutedheteroarlower alkenyl, wherein the substituent(s) is halo, dihalo, loweralkyl, hydroxy, lower alkoxy, amino, aminomethyl, acylamino (acetylamino or benzoylamino) diloweralkylamino, loweralkylamino, carboxyl,haloloweralkyl, cyano or sulfonamido; arloweralkyl or heteroarloweralkylsubstituted on the alkyl portion by amino or acylamino (acetylamino orbenzoylamino);

R² and R⁷ are the same or different and are hydrogen or lower alkyl;

R³ is hydrogen, lower alkyl, phenyl lower alkyl, aminomethyl phenyllower alkyl, hydroxy phenyl lower alkyl, hydroxy lower alkyl, acylaminolower alkyl (such as benzoylamino lower alkyl, acetylamino lower alkyl),amino lower alkyl, dimethylamino lower alkyl, halo lower alkyl,guanidino lower alkyl, imidazolyl lower alkyl, indolyllower alkyl,mercapto lower alkyl, lower alkyl thio lower alkyl;

R⁴ is hydrogen or lower alkyl;

R⁵ is hydrogen, lower alkyl, phenyl, phenyl lower alkyl, hydroxy phenyllower alkyl, hydroxy lower alkyl, amino lower alkyl, guanidino loweralkyl, imidazolyl lower alkyl, indolyl lower alkyl, mercapto lower alkylor lower alkyl thio lower alkyl;

R⁴ and R⁵ can be connected together to form an alkylene bridge of from 2to 4 carbon atoms, an alkylene bridge of from 2 to 3 carbon atoms andone sulfur atom, an alkylene bridge of from 3 to 4 carbon atomscontaining a double bond or an alkylene bridge as above substituted withhydroxy, loweralkoxy, loweralkyl or diloweralky;

-   -   a compound of Formula XIII

wherein R₁ is H, alkyl, acyl or silyl(alkyl)₃; R₂ is H and R₃ is OH,O-acyl, O-alkyl or O-silyl (alkyl)₃ or R₃ is H and R₂ is OH. O-acyl,O-alkyl or O-silyl (alkyl)₃; or R₂ and R₃ together represent O; or R₂and R₃ together represent acetal or cyclic acetal. R₁ might alsorepresent a substituted alkyl such as e.g. methoxy ethoxy methyl;

-   -   a compound of Formula XIV

wherein PY is 4- or 3- or 2-pyridinyl or 4- or 3- or 2-pyridinyl havingone or two lower-alkyl substituents, R is hydrogen, lower-alkyl orlower-hydroxyalkyl, and Q is nitro, carbamyl, halo, amino,lower-alkylamino, di(lower-alkyl)amino, or NHAc where Ac islower-alkanoyl or lower-carbalkoxy;

-   -   a compound of Formula XV

-   -   a compound of Formula XVI

-   -   a compound of Formula XVII

-   -   a compound of Formula XVIII

-   -   a compound of Formula XIX

-   -   a compound of Formula (XX)

or a pharmaceutically acceptable salt thereof.

Further provided is a method of treating or preventing an infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of: Didanosine, Norcyclobenzaprine, Niridazole,Ifosfamide, Cefalonium, Tamoxifen citrate, Butoconazole, Suloctidil,Clomiphene, Sulconazole, Miconazole, Mefloquine, Sulfinpyrazone,Terfenadine, Lisinopril, Econzaole, Clofazimine, Equilin, Felodipine,Dacarbazine, Furazolidone, Perhexiline maleate, Oxethazaine, Pimozide,Trifluoperazine, Ellipticine, Fluspirilen, Hexestrol, Dienestrol,Zidovudine, Metoprolol, Napelline, Methimazole, Amrinone, Iopanoic acid,R-Propanolol, Rimexolone and Pyrvinium pamoate, wherein the infection isa bacterial infection.

Also provided is a method of removing or preventing biofilm formation ona surface, the method comprising administering to a biofilm containingsurface or a surface susceptible to biofilm formation an effectiveamount of a compound selected from the group consisting of:

a compound of Formula I

wherein R represents hydrogen or hydroxyl and R¹ represents hydrogen, orR and R¹ taken together form a second bond between the carbon atomsbearing R and R¹; R² represents hydrogen or phenyl; n is zero or apositive whole integer of from 1 to 4; X represents CH₂, CHOH, NH, C═O,CHNR³R⁴, where R³ and R⁴ independently are hydrogen or lower alkyl; andZ represents thienyl, pyridinyl, substituted pyridinyl, phenyl orsubstituted phenyl wherein the substituents on the substituted pyridinylor substituted phenyl are selected from phenyl, pyridinyl, nitro, ahalogen atom, such as chlorine, fluorine, bromine, or iodine, a straightor branched lower alkyl chain of from 1 to 4 carbon atoms, a loweralkoxy group of from 1 to 4 carbon atoms, amino, a mono ordi(lower)alkylamino group, a saturated monocyclic heterocyclic groupsuch as pyrrolidino, piperidino, morpholino, orN-(lower)alkylpiperazino, or a group having the structure —COOR⁵,—CR⁶R⁷COOR⁵, —CF₃, CHF₂, CH₂F, or

where R⁵ is hydrogen or lower alkyl, and R⁶ and R⁷ independently arehydrogen or methyl or a pharmaceutically acceptable salt thereof.

DESCRIPTION OF DRAWINGS

FIG. 1 depicts the adenylate kinase (AK) assay described in theExamples. Bactericidal agents compromise bacterial cell integrity,releasing cellular adenylate kinase. Extracellular AK is measured by theaddition of a commercial ToxiLight AK cocktail containing ADP andluciferase, resulting in luminescence.

FIG. 2 shows AK assay development. (A) AK assay measures of E. colilysed cell supernatants. E. coli DH5α cells (1×10⁹) were heatinactivated by boiling and diluted, and the AK assay was used to measureadenylate kinase release. Background signal is also shown (mock-treatedcells (right-hand column in FIG. 2A). “*” indicates a significantdifference between results for boiled and mock treatment (Student's ttest; P≦0.05). (B) AK assay results for colistin-treated A. baumanniistrain 98-37-09. Cells were treated with the indicated concentration ofcolistin, and AK was measured; MIC (4 μg ml⁻¹ is indicated. “*”indicates a significant increase in signal compared to that for mocktreatment (0 μg ml⁻¹; Student's t test, P≦0.05).

FIG. 3 shows AK assay measures of S. aureus strain RN4220 treatment withbacteriostatic and bactericidal antibiotics. Standard MIC testingdetermined the MIC of each antibiotic class (in parentheses). Graphedare the fold changes in AK signal of cells treated with 0.5× or 1.0× theMIC value (left hand column and right hand column, respectively) foreach antibiotic, in comparison to untreated control cells; “*” indicatesa significant change in signal as determined by Student's t test; P≦0.05(compared to results for untreated cells).

FIG. 4 shows AK assay measures of antibiotic-treated biofilms andsmall-colony variants. (A) Graphed are AK signals generated by staticbiofilm-associated cells following mock or antibiotic treatment:colistin (P. aeruginosa) or ciprofloxacin (S. aureus and A. baumannii).(B) Fold change of AK measures of S. aureus SCV UAMS-1112 cellsfollowing treatment with 1× and 10× ciprofloxacin, meropenem, andvancomycin, compared to results for mock treated cells. “*” indicates asignificant change in signal in comparison to results for mock-treatedcells (Student's t test, P≦0.05).

FIG. 5 shows AK-based HTS development and screening. (A) Z′ factor assayresults for Klebsiella pneumoniae. Three-hundred-eighty-four-wellmicrotiter plates were seeded with K. pneumoniae, and alternating rowswere mock treated (DMSO) or treated with 50 μM colistin. Following 3 hof incubation, AK release was measured and plotted. DMSO-treated wellmeasures are shown at the bottom of FIG. 5A; colistin-treated wells areshown at the top of FIG. 5A. (B) Prestwick library Klebsiella pneumoniaescreening results. In total, 26 compounds were determined to result in a3-fold increase in AK signal, in comparison to results for DMSO-treatedcells. Included among this list were polymyxin, cephalosporins,aminoglycosides, fluoroquinolones, and detergents; the completePrestwick screening results for K. pneumoniae and all other organismsscreened are provided in Table 4.

FIG. 6 shows antimicrobial properties of terfenadine and tamoxifen. (A)Fold changes in AK signal of terfenadine-treated (10×MIC) S. aureusstrain UAMS-1 static biofilms and the SCV strain UAMS-1112, compared tothose for mock (DMSO)-treated populations, are plotted. “*” indicates asignificant increase in signal over that with mock-treated cells(Student's t test, P≦0.05). (B) Plotted are the percent survival of G.mellonella larvae at 48 h post-E. faecium inoculation. Groups of larvae(n=45) were treated at 2 h and 24 h with either PBS (mock), DMSO, 80 mgkg⁻¹ tamoxifen, 160 mg kg⁻¹ tamoxifen, or 20 mg kg⁻¹ vancomycin.

DETAILED DESCRIPTION

Provided herein is a method of treating or preventing an infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of:

-   -   a compound of Formula I

wherein R represents hydrogen or hydroxyl and R¹ represents hydrogen, orR and R¹ taken together form a second bond between the carbon atomsbearing R and R¹; R² represents hydrogen or phenyl; n is zero or apositive whole integer of from 1 to 4; X represents CH₂, CHOH, NH, C═O,CHNR³R⁴, where R³ and R⁴ independently are hydrogen or lower alkyl; andZ represents thienyl, pyridinyl, substituted pyridinyl, phenyl orsubstituted phenyl wherein the substituents on the substituted pyridinylor substituted phenyl are selected from phenyl, pyridinyl, nitro, ahalogen atom, such as chlorine, fluorine, bromine, or iodine, a straightor branched lower alkyl chain of from 1 to 4 carbon atoms, a loweralkoxy group of from 1 to 4 carbon atoms, amino, a mono ordi(lower)alkylamino group, a saturated monocyclic heterocyclic groupsuch as pyrrolidino, piperidino, morpholino, orN-(lower)alkylpiperazino, or a group having the structure —COOR⁵,—CR⁶R⁷COOR⁵, —CF₃, CHF₂, CH₂F, or

where R⁵ is hydrogen or lower alkyl, and R⁶ and R⁷ independently arehydrogen or methyl. The substituents on the substituted phenyl may beattached at the ortho, meta or para positions of the phenyl ring.

Compounds of Formula I include compounds of Formula II,

wherein R represents hydrogen or hydroxyl and R¹ represents hydrogen, orR and R¹ taken together form a second bond between the carbon atomsbearing R and R¹; n is zero or a positive whole integer of from 1 to 4;Z represents thienyl, pyridinyl, substituted pyridinyl, phenyl orsubstituted phenyl wherein the substituents on the substituted pyridinylor substituted phenyl are selected from phenyl, pyridinyl, nitro, ahalogen atom, such as chlorine, fluorine, bromine, or iodine, a straightor branched lower alkyl chain of from 1 to 4 carbon atoms, a loweralkoxy group of from 1 to 4 carbon atoms, amino, a mono ordi(lower)alkylamino group, a saturated monocyclic heterocyclic groupsuch as pyrrolidino, piperidino, morpholino, orN-(lower)alkylpiperazino, or a group having the structure —COOR⁵,—CR⁶R⁷COOR⁵, —CF₃, CHF₂, CH₂F, or

-   -   where R⁵ is hydrogen or lower alkyl, and R⁶ and R⁷ independently        are hydrogen or methyl    -   or a pharmaceutically acceptable salt thereof.

Compounds of Formula I also include terfenadine and derivatives thereof,including, but not limited to the following compounds. The compounds areidentified by structure, name and registry number. The registry numberfor each compound is also set forth in Table 6 as an additionalidentifier for each compound.

Compound Lot #

Also provided is a method of treating or preventing an infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of:

a compound of Formula III

or a pharmaceutically acceptable salt thereof, wherein R¹ and Rareindependently selected from ethyl or methyl, n 1 or 2, R³ and R⁴ areboth phenyl or substituted phenyl, wherein the substituent can be halo(for example, fluoro-, chloro-, iodo- or bromo-), hydroxyl, a loweralkyl or a substituted lower alkyl wherein the substituent can be halo,hydroxy, or lower alkoxy, and R⁵ is hydrogen, a halogen, a lower alkylfrom about 1 to 4 carbon atoms or a substituted alkyl wherein thesubstituent can be halo, hydroxy, or lower alkoxy. Examples of thecompounds of Formula III include, tamoxifen and derivatives

thereof, including but not limited to tamoxifen (Formula IV), 4-hydroxytamoxifen and clomiphene. In the methods provided herein, wherein a oneor more compounds of Formula III are used to treat or prevent infection,the infection can be an infection, wherein the infection is not a fungalinfection or a parasitic infection.

Further provided herein is a method of treating or preventing aninfection in a subject with or at risk of developing an infection, themethod comprising administering to the subject a compound selected fromthe group consisting of:

-   -   a compound of Formula V

wherein R₁ is a lower alkyl group having from 1 to 4 carbon atoms beingsubstituted with one or several halogen atoms, Z is hydrogen or a loweralkyl group having from 1 to 4 carbon atoms, m is 2 or 3 and X₂ is theethylene imino group or the group having the formula:

wherein R is hydrogen or a lower alkyl group having from 1 to 4 carbonatoms which may be substituted with a chlorine atom or a hydroxy group,and Z and m have the above-given meaning, or a pharmaceuticallyacceptable salt thereof.

As used throughout, the term “lower alkyl group containing from 1 to 4carbon atoms” means methyl, ethyl, n-propyl, isopropyl, n-butyl,isobutyl, tertiary butyl, and I-methylpropyl. The term “halogen” meanschlorine, bromine, fluorine, and iodine. Included herein are compoundsof formula IV that correspond to formula VI:

wherein R₁, X₂ and m have the same meaning as in Formula IV. Additionalcompounds of formula IV, include the compounds of formula VII:

wherein R₂ is a β-chloroethyl or a γ-chloropropyl group, and R₃ ishydrogen, a methyl group or an ethyl group, optionally substituted inthe β-position with a chlorine atom or a hydroxy group. Among thecompounds of formula VII, are the compounds of formula VIII and IX. Thecompound of formula VIII is3-(2-chloroethyl)-2-[(2-chloroethyl)amino]tetrahydro-2H-1,3,2-oxazaphosphorine2-oxide or ifosfamide. Ifosfamide is also known as IFEX.

Further provided herein is a method of treating or preventing aninfection in a subject with or at risk of developing an infection, themethod comprising administering to the subject a compound selected fromthe group consisting of:

a compound of Formula X

wherein R represents a lower alkyl or a lower alkenyl group, a phenylgroup which can be substituted by halogen, methyl or lower alkoxygroups, or a benzyl group which can be nuclear substituted by halogen,methyl or lower alkoxy groups,X represents —O—, —S—, —SO—, or —SO₂—,n represents an integer from 1-4, andAryl represents a phenyl group which can be substituted by lower alkoxyor lower alkylmercapto groups;or a pharmaceutically acceptable salt thereof.

With respect to Formula X, R represents a lower alkyl or a lower alkenylgroup, a phenyl group which can be substituted by halogen, methyl orlower alkoxy groups, or a benzyl group which can be nuclear substitutedby halogen, methyl or lower alkoxy groups, X represents —O—, —S—, —SO—,or —SO2-, n represents an integer from 1-4, and aryl represents a phenylgroup which can be substituted by lower alkoxy or lower alkylmercaptogroups.

For example, —R—X—C_(n)H_(2n)— can represent the following groups:methoxy-, ethoxy-, propoxy-, isopropoxy-, butoxy-, isobutoxy-,allyloxy-, crotyloxy-. phenoxy-, o, -m- and p-methylphenoxy-,o,p-dimethylphenoxy-, m,p-dimethylphenoxy-, p-chlorophenoxy-,p-bromophenoxy-, -o, m and p-methoxyphenoxy-, p-ethoxyphenoxy-,benzyloxy-, o-, m-, and p-methylbenzyloxy-, p-chlorobenzyloxy-,p-bromobenzyloxy, o, m and p-methoxybenzyloxyand-, p-ethoxybenzyloxy-;methyl, -ethyl, -propyl, -isopropyl, and butyl radicals and •analogousradicals with SO or SO₂ instead of O as divalent group X.

Besides the phenyl group, aryl can be, for example, the o- orp-methylmercaptophenyl group, the o- or p-ethyl mercaptophenyl group,the o-; m-, or p-methoxyphenyl group or the o-, m- or p-ethoxyphenylgroup.

An example of the compound of formula X is set forth herein as formulaXI. The compound of formula X is sulfinapyrazone. Sulfinapyrazone isalso known as Anturane.

Also provided herein is a method of treating or preventing an infectionin a subject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of:

a compound of Formula XII

wherein R and R⁶ are the same or different and are hydroxy, loweralkoxy, lower alkenoxy, dilower alkylamino lower alkoxy(dimethylaminoethoxy), acylamino lower alkoxy (acetylaminoethoxy),acyloxy lower alkoxy (pivaloyloxymethoxy), aryloxy, such as phenoxy,arloweralkoxy, such as benzyloxy, substituted aryloxy or substitutedarloweralkoxy wherein the substitutent is methyl, halo or methoxy,amino, loweralkylamino, diloweralkylamino, hydroxyamino,arloweralkylamino such as benzylamino;R¹ is hydrogen, alkyl of from 1 to 20 carbon atoms which includebranched and cyclic and unsaturated (such as allyl) alkyl groups,substituted loweralkyl wherein the substituent can be halo, hydroxy,lower alkoxy, aryloxy such as phenoxy, amino, diloweralkylamino,acylamino, such as acetamido and benzamido, arylamino, guanidino,imidazolyl, indolyl, mercapto, loweralkylthio, arylthio such asphenylthio, carboxy or carboxamido, carboloweralkoxy, aryl such asphenyl or naphthyl, substituted aryl such as phenyl wherein thesubstituent is lower alkyl, lower alkoxy or halo, arloweralkyl,arloweralkenyl, heteroarlower alkyl or heteroarlower alkenyl such asbenzyl, styryl or indolyl ethyl, substituted arloweralkyl, substitutedarloweralkenyl, substituted heteroarlower alkyl, or substitutedheteroarlower alkenyl, wherein the substituent(s) is halo, dihalo, loweralkyl, hydroxy, lower alkoxy, amino, aminomethyl, acylamino (acetylamino or benzoylamino) diloweralkylamino, loweralkylamino, carboxyl,haloloweralkyl, cyano or sulfonamido; arloweralkyl or heteroarloweralkylsubstituted on the alkyl portion by amino or acylamino (acetylamino orbenzoylamino);R² and R⁷ are the same or different and are hydrogen or lower alkyl;R³ is hydrogen, lower alkyl, phenyl lower alkyl, aminomethyl phenyllower alkyl, hydroxy phenyl lower alkyl, hydroxy lower alkyl, acylaminolower alkyl (such as benzoylamino lower alkyl, acetylamino lower alkyl),amino lower alkyl, dimethylamino lower alkyl, halo lower alkyl,guanidino lower alkyl, imidazolyl lower alkyl, indolyllower alkyl,mercapto lower alkyl, lower alkyl thio lower alkyl;R⁴ is hydrogen or lower alkyl;R⁵ is hydrogen, lower alkyl, phenyl, phenyl lower alkyl, hydroxy phenyllower alkyl, hydroxy lower alkyl, amino lower alkyl, guanidino loweralkyl, imidazolyl lower alkyl, indolyl lower alkyl, mercapto lower alkylor lower alkyl thio lower alkyl;R⁴ and R⁵ may be connected together to form an alkylene bridge of from 2to 4 carbon atoms, an alkylene bridge of from 2 to 3 carbon atoms andone sulfur atom, an alkylene bridge of from 3 to 4 carbon atomscontaining a double bond or an alkylene bridge as above substituted withhydroxy, loweralkoxy, loweralkylor diloweralky;or a pharmaceutically acceptable salt thereof.

With respect to Formula XII, R and R⁶ are the same or different and arehydroxy, lower alkoxy, lower alkenoxy, dilower alkylamino lower alkoxy(dimethylaminoethoxy), acylamino lower alkoxy (acetylaminoethoxy),acyloxy lower alkoxy (pivaloyloxymethoxy), aryloxy, such as phenoxy,arloweralkoxy, such as benzyloxy, substituted aryloxy or substitutedarloweralkoxy wherein the substitutent is methyl, halo or methoxy,amino, loweralkylamino, diloweralkylamino, hydroxyamino,arloweralkylamino such as benzylamino;

R¹ is hydrogen, alkyl of from 1 to 20 carbon atoms which includebranched and cyclic and unsaturated (such as allyl) alkyl groups,substituted loweralkyl wherein the substituent can be halo, hydroxy,lower alkoxy, aryloxy such as phenoxy, amino, diloweralkylamino,acylamino, such as acetamido and benzamido, arylamino, guanidino,imidazolyl, indolyl, mercapto, loweralkylthio, arylthio such asphenylthio, carboxy or carboxamido, carboloweralkoxy, aryl such asphenyl or naphthyl, substituted aryl such as phenyl wherein thesubstituent is lower alkyl, lower alkoxy or halo, arloweralkyl,arloweralkenyl, heteroarlower alkyl or heteroarlower alkenyl such asbenzyl, styryl or indolyl ethyl, substituted arloweralkyl, substitutedarloweralkenyl, substituted heteroarlower alkyl, or substitutedheteroarlower alkenyl, wherein the substituent(s) is halo, dihalo, loweralkyl, hydroxy, lower alkoxy, amino, aminomethyl, acylamino (acetylamino or benzoylamino) diloweralkylamino, loweralkylamino, carboxyl,haloloweralkyl, cyano or sulfonamido; arloweralkyl or heteroarloweralkylsubstituted on the alkyl portion by amino or acylamino (acetylamino orbenzoylamino);

R² and R⁷ are the same or different and are hydrogen or lower alkyl;

R³ is hydrogen, lower alkyl, phenyl lower alkyl, aminomethyl phenyllower alkyl, hydroxy phenyl lower alkyl, hydroxy lower alkyl, acylaminolower alkyl (such as benzoylamino lower alkyl, acetylamino lower alkyl),amino lower alkyl, dimethylamino lower alkyl, halo lower alkyl,guanidino lower alkyl, imidazolyl lower alkyl, indolyllower alkyl,mercapto lower alkyl, lower alkyl thio lower alkyl;

R⁴ is hydrogen or lower alkyl;

R⁵ is hydrogen, lower alkyl, phenyl, phenyl lower alkyl, hydroxy phenyllower alkyl, hydroxy lower alkyl, amino lower alkyl, guanidino loweralkyl, imidazolyl lower alkyl, indolyl lower alkyl, mercapto lower alkylor lower alkyl thio lower alkyl;

R⁴ and R⁵ may be connected together to form an alkylene bridge of from 2to 4 carbon atoms, an alkylene bridge of from 2 to 3 carbon atoms andone sulfur atom, an alkylene bridge of from 3 to 4 carbon atomscontaining a double bond or an alkylene bridge as above substituted withhydroxy, loweralkoxy, loweralkylor diloweralky.

The loweralkyl or lower alkenyl groups except where noted otherwiserepresented by any of the variables include straight and branched chainhydrocarbon radicals from one to six carbon atoms, for example, methyl,ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, isopentyl,hexyl or vinyl, allyl, butenyl and the like. The aralkyl groupsrepresented by any of the above variables have from one to four carbonatoms in the alkyl portion thereof and include for example, benzyl,p-methoxy benzyl and the like. Halo means chloro, bromo, iodo or fluoro.Aryl where it appears in any of the radicals except where notedrepresents phenyl or naphthyl. Heteroaryl groups where they appearinclude for example pyridyl, thienyl, furyl, indolyl, benzthienyl,imidazoyl and thiazolyl. The R¹, R³ and R⁵ substituted lower alkylmoieties are exemplified by groups such as

R⁴ and R⁵ when joined through the carbon and nitrogen atoms to whichthey are attached form a 4 to 6 membered ring which may contain onesulfur atom or a double bond. Preferred rings have the formulae:

where Y is CH₂, S, or CHOCH₃.

Compounds of Formula XI include compounds wherein:

R and R⁶ can each independently be hydroxy, lower alkoxy, loweralkenoxy, arloweralkyloxy, amino, dilower alkylamino lower alkoxy,acylamino lower alkoxy, acyloxy lower alkoxy wherein the substituent ismethyl, halo or methoxy;

R² and R⁷ are hydrogen; R³ is lower alkyl, amino lower alkyl,imidazoyllower alkyl, halo lower alkyl;

R⁴ and R⁵ are joined to form the preferred rings as defined above whereY is CH₂, S, or CH—OCH₃;

R¹ is as defined previously.

Other compounds include compounds of Formula XI wherein further R¹ isalkyl having from 1 to 8 carbon atoms, substituted lower alkyl whereinthe alkyl group has 1-5 carbon atoms and the substituent is amino,arylthio, aryloxy or arylamino, aralkyl or heteroaralkyl wherein thealkyl portion has 1 to 3 carbon atoms such as phenethyl or indolylethylor substituted arloweralkyl 65 (phenyl lower alkyl or naphthyl loweralkyl) and substituted heteroarloweralkyl wherein the alkyl groups have1-3 carbons and wherein the substituent(s) is halo, dihalo, amino,aminoalkyl, hydroxy, lower alkoxy or lower alkyl.

Other compounds of Formula XI include compounds wherein R and R⁶ arehydroxy, lower alkoxy, aralkyloxy;

R² and R⁷ are hydrogen;

R³ is methyl or amino lower alkyl;

R⁴ and R⁵ are joined through the carbon and nitrogen atom to formproline, 4-thiaproline or 4-methoxy proline;

R¹ is alkyl having from 1 to 8 carbon atoms, substituted lower alkylwherein the alkyl group has 1-5 carbon atoms and the substituent isamino, arylthio or aryloxy, aralkyl or heteroaralkyl wherein the alkylportion has 1 to 3 carbon atoms such as phenethyl or indolylethyl orsubstituted aralkyl (phenyl lower alkyl or naphthyl lower alkyl) andsubstituted heteroaralkyl wherein the alkyl groups have 1-3 carbons andwherein the substituent(s) is halo, dihalo, amino, aminoalkyl, hydroxy,lower alkoxy or lower alkyl.

Further examples of compounds of Formula XII include, but are notlimited to:

-   N-(1(S)-carboxy-3-phenylpropyl)-L-alanyl-L-proline;-   N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanyl-L-proline and its    maleate salt;-   N-(1(S)-ethoxycarbonyl-4-methylpentyl)-L-alanyl-L-proline;-   N-(1-carboxy-5-aminopentyl)-L-alanyl-L-proline;-   N-.alpha.-(1(S)-carboxy-3-phenylpropyl)-L-lysyl-L-proline    (lisinopril);-   N-.alpha.(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-lysyl-L-proline;-   N-.alpha.[1(S)-carboxy-3-(3-indolyl)propyl]-L-lysyl-L-proline;-   N-.alpha.-[1(S)-carboxy-3-(4-chlorophenyl)-propyl]-L-lysyl-L-proline;-   N-.alpha.-[1(S)-carboxy-2-phenylthioethyl]-L-lysyl-L-proline;-   N-.alpha.-[1(S)-carboxy-3-(4-chlorophenyl)-propyl]-L-lysyl-L-4.alpha.-methoxyproline;-   N-.alpha.-[1(S)-carboxy-5-aminopentyl]-L-lysyl-L-proline;-   Ethyl N-(1(S)-ethoxycarbonyl-3-phenylpropyl)-L-alanyl-L-prolinate    hydrochloride; and the like.

Also provided herein is a method of treating or preventing an infectionin a subject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of:

a compound of Formula XIII

wherein R₁ is H, alkyl, acyl or silyl(alkyl)₃; R₂ is H and R₃ is OH,O-acyl, O-alkyl or O-silyl (alkyl)₃ or R₃ is H and R₂ is OH. O-acyl,O-alkyl or O-silyl (alkyl)₃; or R₂ and R₃ together represent O; or R₂and R₃ together represent acetal or cyclic acetal. R₁ might alsorepresent a substituted alkyl such as e.g. methoxy ethoxy methyl; or apharmaceutically acceptable salt thereof.

Also provided herein is a method of treating or preventing an infectionin a subject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of:

a compound of Formula XIV

wherein PY is 4- or 3- or 2-pyridinyl or 4- or 3- or 2-pyridinyl havingone or two lower-alkyl substituents, R is hydrogen, lower-alkyl orlower-hydroxyalkyl, and Q is nitro, carbamyl, halo, amino,lower-alkylamino, di(lower-alkyl)amino, or NHAc where Ac islower-alkanoyl or lower-carbalkoxy;or a pharmaceutically acceptable salt thereof.

With respect to Formula XIII, PY is 4- or 3- or 2-pyridinyl or 4- or 3-or 2-pyridinyl having one or two lower-alkyl substituents, R ishydrogen, lower-alkyl or lower-hydroxyalkyl, and Q is nitro, carbamyl,halo, amino, lower-alkylamino, di(lower-alkyl)amino, or NHAc where Ac islower-alkanoyl or lower-carbalkoxy, or pharmaceutically-acceptableacid-addition salt thereof. Compounds of formula XIII where Q is amino,lower-alkylamino, di-(lower-alkyl) amino, or NHAc are provided herein.The compounds of formula XIII where Q is nitro or carbamyl are useful asintermediates for preparing the said compounds where Q is amino andthose where Q is halo are useful as intermediates in the preparation ofthe compounds where Q is lower-alkylamino and di-(lower-alkyl)amino.Other compounds of Formula XIII include compounds where Q is amino, R ishydrogen and PY is 4-pyridinyl or 3-pyridinyl, for example,3-amino-5-(4-pyridinyl)-2(1H)-pyridinone (amrinone).

Further provided herein is a method of treating or preventing aninfection in a subject with or at risk of developing an infection, themethod comprising administering to the subject a compound selected fromthe group consisting of:

a compound of Formula XV

a compound of Formula XVI

a compound of Formula XVII

a compound of Formula XVIII

a compound of Formula XIX

a compound of Formula (XX)

or pharmaceutically acceptable salt thereof

Formula XV is fluspirilen, Formula XVI is hexestrol, Formula XVII isdienestrol, Formula XVIII is napelline, Formula XIX is iopanoic acid andFormula XX is suloctodil.

Further provided is a method of treating or preventing an infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of: Didanosine, Norcyclobenzaprine, Niridazole,Ifosfamide, Cefalonium, Tamoxifen citrate, Butoconazole, Suloctidil,Clomiphene, Sulconazole, Miconazole, Mefloquine, Sulfinpyrazone,Terfenadine, Lisinopril, Econzaole, Clofazimine, Equilin, Felodipine,Dacarbazine, Furazolidone, Perhexiline maleate, Oxethazaine, Pimozide,Trifluoperazine, Ellipticine, Fluspirilen, Hexestrol, Dienestrol,Zidovudine, Metoprolol, Napelline, Methimazole, Amrinone, Iopanoic acid,R-Propanolol, Rimexolone and Pyrvinium pamoate, or a pharmaceuticallyacceptable salt thereof.

Further provided is a method of treating or preventing an infection in asubject with or at risk of developing an infection, the methodcomprising administering to the subject a compound selected from thegroup consisting of: Didanosine, Norcyclobenzaprine, Niridazole,Ifosfamide, Cefalonium, Tamoxifen citrate, Butoconazole, Suloctidil,Clomiphene, Sulconazole, Miconazole, Mefloquine, Sulfinpyrazone,Terfenadine, Lisinopril, Econzaole, Clofazimine, Equilin, Felodipine,Dacarbazine, Furazolidone, Perhexiline maleate, Oxethazaine, Pimozide,Trifluoperazine, Ellipticine, Fluspirilen, Hexestrol, Dienestrol,Zidovudine, Metoprolol, Napelline, Methimazole, Amrinone, Iopanoic acid,R-Propanolol, Rimexolone and Pyrvinium pamoate, or a pharmaceuticallyacceptable salt thereof, wherein the infection is a bacterial infectionselected from the group consisting of Enterobacterium faecium,Staphylococcus aureus, Klebsiella pneumonia, Acinebacter baumannii,Pseudomonas aeruginosa and Enterobacter sp.

Further provided is a method of treating or preventing a bacterialinfection in a subject with or at risk of developing a bacterialinfection in a subject comprising administering to the subject acompound that inhibits bacterial DNA gyrase or topoisomerase IV. Thecompound can be, for example, a compound of Formula I. As set forthabove, a compound of Formula I can be terfenadine or a derivativethereof. In the methods set forth herein an inhibitor of bacterial DNAgyrase or topoisomerase IV can be used to treat or preventStaphylococcus aureus infection.

It is contemplated that one or more, for example, two, three, four,five, etc., of the compounds or derivatives of the compounds set forthherein can be administered to treat or prevent infection. Thus,combinations of the compounds set forth herein are also provided.Pharmaceutically acceptable salts of all of the compounds set forthherein are also provided. The term pharmaceutically acceptable salt asused herein refers to those salts of any of the compounds describedherein or derivatives thereof that are, within the scope of soundmedical judgment, suitable for use in contact with the tissues ofsubjects without undue toxicity, irritation, allergic response, and thelike, commensurate with a reasonable benefit/risk ratio, and effectivefor their intended use, as well as the zwitterionic forms, wherepossible, of the compounds described herein. The term salts refers tothe relatively non-toxic, inorganic and organic acid addition salts ofthe compounds described herein. These salts can be prepared in situduring the isolation and purification of the compounds or by separatelyreacting the purified compound in its free base form with a suitableorganic or inorganic acid and isolating the salt thus formed.Representative salts include the hydrobromide, hydrochloride, sulfate,bisulfate, nitrate, acetate, oxalate, valerate, oleate, palmitate,stearate, laurate, borate, benzoate, lactate, phosphate, tosylate,citrate, maleate, fumarate, succinate, tartrate, naphthylate mesylate,glucoheptonate, lactobionate, methane sulphonate, and laurylsulphonatesalts, and the like. These may include cations based on the alkali andalkaline earth metals, such as sodium, lithium, potassium, calcium,magnesium, and the like, as well as non-toxic ammonium, quaternaryammonium, and amine cations including, but not limited to ammonium,tetramethylammonium, tetraethylammonium, methylamine, dimethylamine,trimethylamine, triethylamine, ethylamine, and the like.

The infection can be a viral infection, bacterial infection, fungalinfection or a parasitic infection, to name a few. All strains and typesof pathogenic infection are contemplated herein. The infection can alsobe a respiratory infection, a gastrointestinal infection or a skininfection, to name a few.

In any of the methods of treating or preventing infection set forthherein, the infection can be any infection, wherein the infection is nota bacterial infection. In any of the methods of treating or preventinginfection set forth herein, the infection can be any infection, whereinthe infection is not a viral infection. In any of the methods oftreating or preventing infection set forth herein, the infection can beany infection, wherein the infection is not a parasitic infection. Inany of the methods of treating or preventing infection set forth herein,the infection can be any infection, wherein the infection is not afungal infection. In any of the methods of treating or preventinginfection set forth herein, the infection can be any infection, whereinthe infection is not a protozoal infection.

Examples of bacterial infections include, but are not limited toinfections caused by the Gram negative or Gram positive bacteria. Forexample, the infection can be caused by Listeria (sp.), Franscicellatularensis, Enterobacter sp. Enterococcus faecium, other Enterococcusspecies, Klebsiella pneumonia, Acinetobacter baumannii, Mycobacteriumtuberculosis, Rickettsia (all types), Ehrlichia or Chylamida. Furtherexamples of bacteria include M. tuberculosis, Legionella pneumophila,other Legionella species, Salmonella typhi, other Salmonella species,Shigella species, Yersinia pestis, Pasteurella haemolytica, Pasteurellamultocida, other Pasteurella species, Actinobacillus pleuropneumoniae,Listeria monocytogenes, Listeria ivanovii, Brucella abortus, otherBrucella species, Chlamydia pneumoniae, Chlamydia trachomatis, Chlamydiapsittaci, Coxiella burnetti, other Rickettsial species, Ehrlichiaspecies, Staphylococcus aureus, Staphylococcus epidermidis,Streptococcus pyogenes, Streptococcus agalactiae, Bacillus anthracis,Escherichia coli, Vibrio cholerae, Kingella kingae, Campylobacterspecies, Neiserria meningitidis, Neiserria gonorrhea, Pseudomonasaeruginosa, other Pseudomonas species, Haemophilus influenzae,Haemophilus ducreyi, other Hemophilus species, Clostridium tetani, otherClostridium species, Yersinia enterolitica, and other Yersinia species.In the methods provided herein, one or more compounds set forth hereincan treat or prevent one or more bacterial infections selected from thegroup consisting of Enterobacterium faecium, Staphylococcus aureus,Klebsiella pneumonia, Acinebacter baumannii, Pseudomonas aeruginosa andEnterobacter sp. In the methods set forth herein, the bacteria can be asmall colony variant strain, for example a small colony strain ofStaphylococcus aureus. In any of the methods set forth herein, theinfection can be a bacterial infection, wherein the bacterial infectionis not tuberculosis, for example, Mycobacterium tuberculosis. Forexample, and not to be limiting any of the compounds set forth herein,including compounds of Formula I and II can be used to treat or preventinfection in a subject with or at risk of developing an infection,wherein the infection is not tuberculosis.

Examples of parasitic infections include, but are not limited toinfections caused by the following parasites: Cryptosporidium,Plasmodium (all species), American trypanosomes (T. cruzi), Africantrypanosomes, Acanthamoeba, Entaoeba histolytica, Angiostrongylus,Anisakis, Ascaris, Babesia, Balantidium, Baylisascaris, lice, ticks,mites, fleas, Capillaria, Clonorchis, Chilomastix mesnili, Cyclspora,Diphyllobothrium, Dipylidium caninum, Fasciola, Giardia, Gnathostoma,Hetetophyes, Hymenolepsis, Isospora, Loa loa, Microsporidia, Naegleria,Toxocara, Onchocerca, Opisthorchis, Paragonimus, Baylisascaris,Strongyloides, Taenia, Trichomonas and Trichuris. In any of the methodsset forth herein, the infection can be a parasitic infection, whereinthe parasitic infection is not malaria, for example, malaria caused byany species of Plasmodium including Plasmodium falciparum. For example,and not to be limiting any of the compounds set forth herein, includingcompounds of Formula I and II can be used to treat or prevent infectionin a subject with or at risk of developing an infection, wherein theinfection is not malaria.

Furthermore, examples of protozoan and fungal species contemplatedwithin the present methods include, but are not limited to, Plasmodiumfalciparum, other Plasmodium species, Toxoplasma gondii, Pneumocystiscarinii, Trypanosoma cruzi, other trypanosomal species, Leishmaniadonovani, other Leishmania species, Theileria annulata, other Theileriaspecies, Eimeria tenella, other Eimeria species, Histoplasma capsulatum,Cryptococcus neoformans, Blastomyces dermatitidis, Coccidioides immitis,Paracoccidioides brasibiensis, Penicillium marneffei, and Candidaspecies. In any of the methods set forth herein, the infection can be aprotozoan infection, wherein the protozoan infection is notleishmaniasis, for example, leishmaniasis caused by a Leishmaniaspecies, for example, Leishmania major. For example, and not to belimiting any of the compounds set forth herein, including compounds ofFormula I and III can be used to treat or prevent infection in a subjectwith or at risk of developing an infection, wherein the infection is notleishmaniasis.

Examples of viral infections include but are not limited to, infectionscaused by RNA viruses (including negative stranded RNA viruses, positivestranded RNA viruses, double stranded RNA viruses and retroviruses) andDNA viruses. All strains, types, subtypes of DNA and RNA viruses arecontemplated herein.

Examples of RNA viruses include, but are not limited to picornaviruses,which include aphthoviruses (for example, foot and mouth disease virusO, A, C, Asia 1, SAT1, SAT2 and SAT3), cardioviruses (for example,encephalomycarditis virus and Theiller's murine encephalomyelitisvirus), enteroviruses (for example polioviruses 1, 2 and 3, humanenteroviruses A-D, bovine enteroviruses 1 and 2, human coxsackievirusesA1-A22 and A24, human coxsackieviruses B1-B5, human echoviruses 1-7, 9,11-12, 24, 27, 29-33, human enteroviruses 68-71, porcine enteroviruses8-10 and simian enteroviruses 1-18), erboviruses (for example, equinerhinitis virus), hepatovirus (for example human hepatitis A virus andsimian hepatitis A virus), kobuviruses (for example, bovine kobuvirusand Aichi virus), parechoviruses (for example, human parechovirus 1 andhuman parechovirus 2), rhinovirus (for example, human rhinovirus 1-100and bovine rhinoviruses 1-3) and teschoviruses (for example, porcineteschovirus).

Additional examples of RNA viruses include caliciviruses, which includenoroviruses (for example, Norwalk virus), sapoviruses (for example,Sapporo virus), lagoviruses (for example, rabbit hemorrhagic diseasevirus and European brown hare syndrome) and vesiviruses (for examplevesicular exanthema of swine virus and feline calicivirus).

Other RNA viruses include astroviruses, which include mamastorvirusesand avastroviruses. Togaviruses are also RNA viruses. Togavirusesinclude alphaviruses (for example, Chikungunya virus, Sindbis virus,Semliki Forest virus, Western equine encephalitis, Getah virus,Everglades virus, Venezuelan equine encephalitis virus and Aura virus)and rubella viruses. Additional examples of RNA viruses include the theflaviviruses (for example, tick-borne encephalitis virus, Tyuleniyvirus, Aroa virus, Dengue virus (types 1 to 4), Kedougou virus, Japaneseencephalitis virus (JEV), West Nile virus (WNV), Kokobera virus, Ntayavirus, Spondweni virus, Yellow fever virus, Entebbe bat virus, Modocvirus, Rio Bravo virus, Cell fusing agent virus, pestivirus, GB virus A,GBV-A like viruses, GB virus C, Hepatitis G virus, hepacivirus(hepatitis C virus (HCV)) all six genotypes), bovine viral diarrheavirus (BVDV) types 1 and 2, and GB virus B).

Other examples of RNA viruses are the coronaviruses, which include,human respiratory coronaviruses such as SARS-CoV, HCoV-229E, HCoV-NL63and HCoV-OC43. Coronaviruses also include bat SARS-like CoV, turkeycoronavirus, chicken coronavirus, feline coronavirus and caninecoronavirus. Additional RNA viruses include arteriviruses (for example,equine arterivirus, porcine reproductive and respiratory syndrome virus,lactate dehyrogenase elevating virus of mice and simian hemorraghicfever virus). Other RNA viruses include the rhabdoviruses, which includelyssaviruses (for example, rabies, Lagos bat virus, Mokola virus,Duvenhage virus and European bat lyssavirus), vesiculoviruses (forexample, VSV-Indiana, VSV-New Jersey, VSV-Alagoas, Piry virus, Cocalvirus, Maraba virus, Isfahan virus and Chandipura virus), andephemeroviruses (for example, bovine ephemeral fever virus, AdelaideRiver virus and Berrimah virus). Additional examples of RNA virusesinclude the filoviruses. These include the Marburg and Ebola viruses(for example, EBOV-Z, EBOV-S, EBOV-IC and EBOV-R.

The paramyxoviruses are also RNA viruses. Examples of these viruses arethe rubulaviruses (for example, mumps, parainfluenza virus 5, humanparainfluenza virus type 2, Mapuera virus and porcine rubulavirus),avulaviruses (for example, Newcastle disease virus), respoviruses (forexample, Sendai virus, human parainfluenza virus type 1 and type 3,bovine parainfluenza virus type 3), henipaviruses (for example, Hendravirus and Nipah virus), morbilloviruses (for example, measles, Cetaceanmorvilliirus, Canine distemper virus, Peste-des-petits-ruminants virus,Phocine distemper virus and Rinderpest virus), pneumoviruses (forexample, human respiratory syncytial virus A2, B1 and S2, bovinerespiratory syncytial virus and pneumonia virus of mice),metapneumoviruses (for example, human metapneumovirus and avianmetapneumovirus). Additional paramyxoviruses include Fer-de-Lance virus,Tupaia paramyxovirus, Menangle virus, Tioman virus, Beilong virus, Jvirus, Mossman virus, Salem virus and Nariva virus. Additional RNAviruses include the orthomyxoviruses.

These viruses include influenza viruses and strains (e.g., influenza A(H1N1 (including but not limited to A/WS/33 and A/California/04/2009strains) H2N2, H3N2, H5N1, H7N7, H1N2, H9N2, H7N2, H7N3 and H10N7), Band C viruses, as well as avian influenza (for example, strains H5N1,H5N2, H7N1, H7N7 and H9N2) thogotoviruses and isaviruses.Orthobunyaviruses (for example, Akabane virus, California encephalitis,Cache Valley virus, Snowshoe hare virus,) nairoviruses (for example,Nairobi sheep virus, Crimean-Congo hemorrhagic fever virus Group andHughes virus), phleboviruses (for example, Candiru, Punta Toro, RiftValley Fever, Sandfly Fever, Naples, Toscana, Sicilian and Chagres), andhantaviruses (for example, Hantaan, Dobrava, Seoul, Puumala, Sin Nombre,Bayou, Black Creek Canal, Andes and Thottapalayam) are also RNA viruses.Arenaviruses such as lymphocytic choriomeningitis virus, Lujo virus,Lassa fever virus, Argentine hemorrhagic fever virus, Bolivianhemorrhagic fever virus, Venezuelan hemorrhagic fever virus, SABV andWWAV are also RNA viruses. Borna disease virus is also an RNA virus.Hepatitis D (Delta) virus and hepatitis E are also RNA viruses. Any ofthe compounds set forth herein, including, but not limited to thecompounds of Formula I and II can be used to treat or prevent a viralinfection, wherein the viral infection is not a Lassa fever virusinfection.

Additional RNA viruses include reoviruses, rotaviruses, birnaviruses,chrysoviruses, cystoviruses, hypoviruses partitiviruses and totoviruses.Orbiviruses such as African horse sickness virus, Blue tongue virus,Changuinola virus, Chenuda virus, Chobar Gorge Corriparta virus,epizootic hemorraghic disease virus, equine encephalosis virus,Eubenangee virus, Ieri virus, Great Island virus, Lebombo virus, Orungovirus, Palyam virus, Peruvian Horse Sickness virus, St. Croix Rivervirus, Umatilla virus, Wad Medani virus, Wallal virus, Warrego virus andWongorr virus are also RNA viruses.

Retroviruses include alpharetroviruses (for example, Rous sarcoma virusand avian leukemia virus), betaretroviruses (for example, mouse mammarytumor virus, Mason-Pfizer monkey virus and Jaagsiekte sheep retrovirus),gammaretroviruses (for example, murine leukemia virus and felineleukemia virus, deltraretroviruses (for example, human T cell leukemiaviruses (HTLV-1, HTLV-2), bovine leukemia virus, STLV-1 and STLV-2),epsilonretriviruses (for example, Walleye dermal sarcoma virus andWalleye epidermal hyperplasia virus 1), reticuloendotheliosis virus (forexample, chicken syncytial virus, lentiviruses (for example, humanimmunodeficiency virus (HIV) type 1, human immunodeficiency virus (HIV)type 2, human immunodeficiency virus (HIV) type 3, simianimmunodeficiency virus, equine infectious anemia virus, felineimmunodeficiency virus, caprine arthritis encephalitis virus and Visnamaedi virus) and spumaviruses (for example, human foamy virus and felinesyncytia-forming virus).

Examples of DNA viruses include polyomaviruses (for example, simianvirus 40, simian agent 12, BK virus, JC virus, Merkel Cell polyomavirus, bovine polyoma virus and lymphotrophic papovavirus),papillomaviruses (for example, human papillomavirus, bovinepapillomavirus, adenoviruses (for example, adenoviruses A-F, canineadenovirus type I, canined adeovirus type 2), circoviruses (for example,porcine circovirus and beak and feather disease virus (BFDV)),parvoviruses (for example, canine parvovirus), erythroviruses (forexample, adeno-associated virus types 1-8), betaparvoviruses,amdoviruses, densoviruses, iteraviruses, brevidensoviruses,pefudensoviruses, herpes viruses 1, 2, 3, 4, 5, 6, 7 and 8 (for example,herpes simplex virus 1, herpes simplex virus 2, varicella-zoster virus,Epstein-Barr virus, cytomegalovirus, Kaposi's sarcoma associated herpesvirus, human herpes virus-6 variant A, human herpes virus-6 variant Band cercophithecine herpes virus 1 (B virus)), poxviruses (for example,smallpox (variola), cowpox, monkeypox, vaccinia, Uasin Gishu, camelpox,psuedocowpox, pigeonpox, horsepox, fowlpox, turkeypox and swinepox), andhepadnaviruses (for example, hepatitis B and hepatitis B-like viruses).

One or more of the compounds described herein can be contacted with acell or populations of cells in vitro, ex vivo or in vivo. For example,the cell or population of cells can be in a subject, or in an in vitroculture. In another example, one or more compounds set forth herein canbe used to inhibit bacterial growth, fungal growth, parasitic growth,protozoal growth or viral replication, in vitro, ex vivo or in vivo. Anyof the compounds set forth herein can be used alone or in combinationwith other therapeutic agents such as antiviral compounds, antibacterialagents (for example, antibiotics), antifungal agents, antiparasiticagents, anti-inflammatory agents, anti-cancer agents, etc.

In the methods described herein, the level of infection, for example, ina cell, or a population of cells, or a cell culture, can be assessed bymeasuring an antigen or other product associated with a particularinfection. The level of infection can also be measured in a tissuesample or a culture of cells from a subject, either before or afteradministration of one or more compounds disclosed herein. For example,the level of viral infection can be measured by real-time quantitativereverse transcription-polymerase chain reaction (RT-PCR) assay (See forexample, Payungporn et al. “Single step multiplex real-time RT-PCR forH5N1 influenza A virus detection.” J Virol Methods. Sep. 22, 2005;Landolt et la. “Use of real-time reverse transcriptase polymerase chainreaction assay and cell culture methods for detection of swine influenzaA viruses” Am J Vet Res. 2005 January; 66(1):119-24).

Methods of measuring bacterial growth and inhibition of bacterial growthare provided in the Examples. Further, one of skill in the art wouldknow how to determine the concentration of a compound that inhibitsbacterial infection (see, for example, Andrews, et al. “Determination ofminimum inhibitory concentrations”. Journal of AntimicrobialChemotherapy 48 (suppl 1): 5-16 (2001)). Other methods for determiningantifungal and antibacterial activity are known in the art. See, forexample, Hayhoe et al. “Screening for Antibacterial, Antifungal and Antiquorum Sensing Activity,” Methods Mol. Biol. 1055: 219-225 (2013));Doddanna et al. “Antimicrobial activity of plant extracts on Candidaalbicans: An in vitro study Indian J. Dent. Res. 24(4): 401-405 (2013),both of which are incorporated by this reference in their entireties.

As used throughout, by subject is meant an individual. Preferably, thesubject is a mammal such as a primate, and, more preferably, a human.Non-human primates are subjects as well. The term subject includesdomesticated animals, such as cats, dogs, etc., livestock (for example,cattle, horses, pigs, sheep, goats, etc.) and laboratory animals (forexample, ferret, chinchilla, mouse, rabbit, rat, gerbil, guinea pig,etc.). Thus, veterinary uses and medical formulations are contemplatedherein.

As used herein, a biological sample is a sample derived from a subjectsuch as a mammal or human and includes, but is not limited to, anybiological fluid, including a bodily fluid. Examples of bodily fluidsinclude, but are not limited to, whole blood, plasma, serum, urine,saliva, ocular fluid, ascites, a stool sample, spinal fluid, tissueinfiltrate, pleural effusions, lung lavage fluid, and the like. Thebiological fluid includes a cell culture medium or supernatant ofcultured cells from the subject.

The methods and compounds as described herein are useful for therapeutictreatment. Use of one or more of the compounds set forth herein for thetreatment or prevention of infection is also contemplated herein. One ormore of the compounds set forth herein for use in a method of treatingor preventing infection is also provided herein. Therapeutic treatmentinvolves administering to a subject a therapeutically effective amountof one or more of the agents described herein, optionally, afterdiagnosis of an infection or risk of infection in the subject.Therefore, all of the methods disclosed herein, can optionally comprisethe step of diagnosing a subject with an infection or diagnosing asubject in need of prophylaxis or prevention of infection.

As used herein, the terms treatment, treat, or treating refers to amethod of reducing the effects of a disease or condition or symptom ofthe disease or condition. Thus, in the disclosed methods, treatment canrefer to a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%reduction in the severity of an established disease or condition orsymptom of the disease or condition. For example, a method for treatinga disease is considered to be a treatment if there is a 10% reduction inone or more symptoms of the disease in a subject as compared to acontrol. A control subject can be a subject that has not received acompound set forth herein. Thus, the reduction can be a 10%, 20%, 30%,40%, 50%, 60%, 70%, 80%, 90%, 100%, or any percent reduction in between10% and 100% as compared to native or control levels. It is understoodthat treatment does not necessarily refer to a cure or complete ablationof the disease, condition, or symptoms of the disease or condition.

As utilized herein, by preventing infection is meant a method ofprecluding, delaying, averting, obviating, forestalling, stopping, orhindering the onset, incidence, severity, or recurrence of infection.For example, the disclosed method is considered to be a prevention ifthere is about a 10% reduction in onset, incidence, severity, orrecurrence of infection, or symptoms of infection (e.g., inflammation,fever, lesions, weight loss, etc.) in a subject exposed to an infectionwhen compared to control subjects exposed to an infection that did notreceive a composition for decreasing infection. Thus, the reduction inonset, incidence, severity, or recurrence of infection can be about a10, 20, 30, 40, 50, 60, 70, 80, 90, 100%, or any amount of reduction inbetween as compared to a control subject. For example, and not to belimiting, if about 10% of the subjects in a population do not becomeinfected as compared to subjects that did not receive preventivetreatment, this is considered prevention.

The compounds set forth herein can also be used to decrease infection ina cell. A decrease or inhibition of infection can occur in a cell, invitro, ex vivo or in vivo. As utilized throughout, the term “infection”encompasses all phases of pathogenic life cycles including, but notlimited to, attachment to cellular receptors, entry, internalization,disassembly, replication, genomic integration of pathogenic sequences,transcription of pathogen RNA, translation of pathogen RNA,transcription of host cell mRNA, translation of host cell mRNA,proteolytic cleavage of pathogenic proteins or cellular proteins,assembly of particles, endocytosis, cell lysis, budding, and egress ofthe pathogen from the cells.

The compounds described herein can be provided in a pharmaceuticalcomposition. Depending on the intended mode of administration, thepharmaceutical composition can be in the form of solid, semi-solid orliquid dosage forms, such as, for example, tablets, suppositories,pills, capsules, powders, liquids, or suspensions, preferably in unitdosage form suitable for single administration of a precise dosage. Thecompositions will include a therapeutically effective amount of thecompound described herein or derivatives thereof in combination with apharmaceutically acceptable carrier and, in addition, may include othermedicinal agents, pharmaceutical agents, carriers, or diluents. Bypharmaceutically acceptable is meant a material that is not biologicallyor otherwise undesirable, which can be administered to an individualalong with the selected agent without causing unacceptable biologicaleffects or interacting in a deleterious manner with the other componentsof the pharmaceutical composition in which it is contained.

As used herein, the term carrier encompasses any excipient, diluent,filler, salt, buffer, stabilizer, solubilizer, lipid, stabilizer, orother material well known in the art for use in pharmaceuticalformulations. The choice of a carrier for use in a composition willdepend upon the intended route of administration for the composition.The preparation of pharmaceutically acceptable carriers and formulationscontaining these materials is described in, e.g., Remington'sPharmaceutical Sciences, 21st Edition, ed. University of the Sciences inPhiladelphia, Lippincott, Williams & Wilkins, Philadelphia Pa., 2005.Examples of physiologically acceptable carriers include buffers such asphosphate buffers, citrate buffer, and buffers with other organic acids;antioxidants including ascorbic acid; low molecular weight (less thanabout 10 residues) polypeptides; proteins, such as serum albumin,gelatin, or immunoglobulins; hydrophilic polymers such aspolyvinylpyrrolidone; amino acids such as glycine, glutamine,asparagine, arginine or lysine; monosaccharides, disaccharides, andother carbohydrates including glucose, mannose, or dextrins; chelatingagents such as EDTA; sugar alcohols such as mannitol or sorbitol;salt-forming counterions such as sodium; and/or nonionic surfactantssuch as TWEEN® (ICI, Inc.; Bridgewater, N.J.), polyethylene glycol(PEG), and PLURONICS™ (BASF; Florham Park, N.J.).

Compositions containing the compound(s) described herein suitable forparenteral injection may comprise physiologically acceptable sterileaqueous or nonaqueous solutions, dispersions, suspensions or emulsions,and sterile powders for reconstitution into sterile injectable solutionsor dispersions. Examples of suitable aqueous and nonaqueous carriers,diluents, solvents or vehicles include water, ethanol, polyols(propyleneglycol, polyethyleneglycol, glycerol, and the like), suitablemixtures thereof, vegetable oils (such as olive oil) and injectableorganic esters such as ethyl oleate. Proper fluidity can be maintained,for example, by the use of a coating such as lecithin, by themaintenance of the required particle size in the case of dispersions andby the use of surfactants.

These compositions may also contain adjuvants such as preserving,wetting, emulsifying, and dispensing agents. Prevention of the action ofmicroorganisms can be promoted by various antibacterial and antifungalagents, for example, parabens, chlorobutanol, phenol, sorbic acid, andthe like. Isotonic agents, for example, sugars, sodium chloride, and thelike may also be included. Prolonged absorption of the injectablepharmaceutical form can be brought about by the use of agents delayingabsorption, for example, aluminum monostearate and gelatin.

Solid dosage forms for oral administration of the compounds describedherein or derivatives thereof include capsules, tablets, pills, powders,and granules. In such solid dosage forms, the compounds described hereinor derivatives thereof is admixed with at least one inert customaryexcipient (or carrier) such as sodium citrate or dicalcium phosphate or(a) fillers or extenders, as for example, starches, lactose, sucrose,glucose, mannitol, and silicic acid, (b) binders, as for example,carboxymethylcellulose, alignates, gelatin, polyvinylpyrrolidone,sucrose, and acacia, (c) humectants, as for example, glycerol, (d)disintegrating agents, as for example, agar-agar, calcium carbonate,potato or tapioca starch, alginic acid, certain complex silicates, andsodium carbonate, (e) solution retarders, as for example, paraffin, (f)absorption accelerators, as for example, quaternary ammonium compounds,(g) wetting agents, as for example, cetyl alcohol, and glycerolmonostearate, (h) adsorbents, as for example, kaolin and bentonite, and(i) lubricants, as for example, talc, calcium stearate, magnesiumstearate, solid polyethylene glycols, sodium lauryl sulfate, or mixturesthereof. In the case of capsules, tablets, and pills, the dosage formsmay also comprise buffering agents.

Solid compositions of a similar type may also be employed as fillers insoft and hard-filled gelatin capsules using such excipients as lactoseor milk sugar as well as high molecular weight polyethyleneglycols, andthe like.

Solid dosage forms such as tablets, dragees, capsules, pills, andgranules can be prepared with coatings and shells, such as entericcoatings and others known in the art. They may contain opacifying agentsand can also be of such composition that they release the activecompound or compounds in a certain part of the intestinal tract in adelayed manner.

Examples of embedding compositions that can be used are polymericsubstances and waxes. The active compounds can also be inmicro-encapsulated form, if appropriate, with one or more of theabove-mentioned excipients.

Liquid dosage forms for oral administration of the compounds describedherein or derivatives thereof include pharmaceutically acceptableemulsions, solutions, suspensions, syrups, and elixirs. In addition tothe active compounds, the liquid dosage forms may contain inert diluentscommonly used in the art, such as water or other solvents, solubilizingagents, and emulsifiers, as for example, ethyl alcohol, isopropylalcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzylbenzoate, propyleneglycol, 1,3-butyleneglycol, dimethylformamide, oils,in particular, cottonseed oil, groundnut oil, corn germ oil, olive oil,castor oil, sesame oil, glycerol, tetrahydrofurfuryl alcohol,polyethyleneglycols, and fatty acid esters of sorbitan, or mixtures ofthese substances, and the like.

Besides such inert diluents, the composition can also include additionalagents, such as wetting, emulsifying, suspending, sweetening, flavoring,or perfuming agents.

Administration can be carried out using therapeutically effectiveamounts of the agents described herein for periods of time effective totreat or prevent infection in a subject. The effective amount may bedetermined by one of ordinary skill in the art and includes exemplarydosage amounts for a mammal of from about 0.5 to about 200 mg/kg of bodyweight of active compound per day, which may be administered in a singledose or in the form of individual divided doses, such as from 1 to 4times per day. Alternatively, the dosage amount can be from about 0.5 toabout 150 mg/kg of body weight of active compound per day, about 0.5 to100 mg/kg of body weight of active compound per day, about 0.5 to about75 mg/kg of body weight of active compound per day, about 0.5 to about50 mg/kg of body weight of active compound per day, about 0.5 to about25 mg/kg of body weight of active compound per day, about 1 to about 20mg/kg of body weight of active compound per day, about 1 to about 10mg/kg of body weight of active compound per day, about 20 mg/kg of bodyweight of active compound per day, about 10 mg/kg of body weight ofactive compound per day, or about 5 mg/kg of body weight of activecompound per day.

According to the methods taught herein, the subject is administered aneffective amount of the compound. The terms effective amount andeffective dosage are used interchangeably. The term effective amount isdefined as any amount necessary to produce a desired physiologicresponse. Effective amounts and schedules for administering the agentmay be determined empirically, and making such determinations is withinthe skill in the art. The dosage ranges for administration are thoselarge enough to produce the desired effect in which one or more symptomsof the disease or disorder are affected (e.g., reduced or delayed). Thedosage should not be so large as to cause substantial adverse sideeffects, such as unwanted cross-reactions, anaphylactic reactions, andthe like. Generally, the dosage will vary with the activity of thespecific compound employed, the metabolic stability and length of actionof that compound, the species, age, body weight, general health, sex anddiet of the subject, the mode and time of administration, rate ofexcretion, drug combination, and severity of the particular conditionand can be determined by one of skill in the art. The dosage can beadjusted by the individual physician in the event of anycontraindications. Dosages can vary, and can be administered in one ormore dose administrations daily, for one or several days. Guidance canbe found in the literature for appropriate dosages for given classes ofpharmaceutical products.

Any appropriate route of administration can be employed, depending onwhether local or systemic treatment is desired, and on the area to betreated. The compositions are administered via any of several routes ofadministration, including topically, orally, parenterally,intravenously, intra-articularly, intraperitoneally, intramuscularly,subcutaneously, intracavity, transdermally, intrahepatically,intracranially, nebulization/inhalation, or by installation viabronchoscopy. Optionally, the composition is administered by oralinhalation, nasal inhalation, or intranasal mucosal administration.Administration of the compositions by inhalant can be through the noseor mouth via delivery by spraying or droplet mechanism, for example, inthe form of an aerosol. Pharmaceutical compositions can be deliveredlocally to the area in need of treatment, for example by topicalapplication or local injection. Multiple administrations and/or dosagescan also be used. Effective doses can be extrapolated from dose-responsecurves derived from in vitro or animal model test systems.

The disclosure also provides a pharmaceutical pack or kit comprising oneor more containers filled with one or more of the ingredients of thepharmaceutical compositions. Optionally associated with suchcontainer(s) can be a notice in the form prescribed by a governmentalagency regulating the manufacture, use or sale of pharmaceuticals orbiological products, which notice reflects approval by the agency ofmanufacture, use or sale for human administration. Instructions for useof the composition can also be included.

Also provided is a method of removing biofilm from a surface, comprisingadministering an effective amount of one or more of the compoundsprovided herein to a biofilm-containing surface, wherein the amount iseffective to remove biofilm from the surface. Removal of the biofilmfrom this surface does not have to be complete as this can range from areduction to complete removal of the biofilm. Thus, in the disclosedmethods, removal can refer to a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%,90%, or 100% reduction in the amount of biofilm on a surface. Forexample, a method for removing biofilm from a surface is considered tobe removal if there is a 10% reduction in the amount of biofilm on thesurface as compared to a control. A control surface can be a biofilmcontaining surface that has not received a compound set forth herein.Thus, the reduction can be a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%,90%, 100%, or any percent reduction in between 10% and 100% as comparedto control.

Further provided is a method of preventing biofilm formation on asurface comprising administering an effective amount of one or more ofthe compounds provided herein to the surface, wherein the amount iseffective to prevent biofilm formation. The surface can be susceptibleto biofilm formation. The biofilm can be produced by an organismselected from the group consisting of bacteria, algae, fungi andprotozoa.

The compound can be, but is not limited to, a compound of Formula I

wherein R represents hydrogen or hydroxyl and R¹ represents hydrogen, orR and R¹ taken together form a second bond between the carbon atomsbearing R and R¹; R² represents hydrogen or phenyl; n is zero or apositive whole integer of from 1 to 4; X represents CH₂, CHOH, NH, C═O,CHNR³R⁴, where R³ and R⁴ independently are hydrogen or lower alkyl; andZ represents thienyl, pyridinyl, substituted pyridinyl, phenyl orsubstituted phenyl wherein the substituents on the substituted pyridinylor substituted phenyl are selected from phenyl, pyridinyl, nitro, ahalogen atom, such as chlorine, fluorine, bromine, or iodine, a straightor branched lower alkyl chain of from 1 to 4 carbon atoms, a loweralkoxy group of from 1 to 4 carbon atoms, amino, a mono ordi(lower)alkylamino group, a saturated monocyclic heterocyclic groupsuch as pyrrolidino, piperidino, morpholino, orN-(lower)alkylpiperazino, or a group having the structure —COOR⁵,—CR⁶R⁷COOR⁵, —CF₃, CHF₂, CH₂F, or

where R⁵ is hydrogen or lower alkyl, and R⁶ and R⁷ independently arehydrogen or methylor a pharmaceutically acceptable salt thereof.

One or more of the compounds set forth herein can be combined with oneor more biodegradable polymers to form a biodegradable antimicrobialcomposition. These compositions can be applied to a surface or used as acoating. The biodegradable polymers include but are not limited topolylactic acid, polyglycolic acid and copolymers and mixtures thereofsuch as poly(L-lactide) (PLLA), poly(D,L-lactide) (PLA), polyglycolicacid [polyglycolide (PGA)], poly(L-lactide-co-D,L-lactide) (PLLA/PLA),poly(L-lactide-co-glycolide) (PLLA/PGA), poly(D,L-lactide-co-glycolide)(PLA/PGA), poly(glycolide-co-trimethylene carbonate) (PGA/PTMC),poly(D,L-lactide-co-caprolactone) (PLA/PCL) andpoly(glycolide-co-caprolactone) (PGA/PCL); polyethylene oxide (PEO),polydioxanone (PDS), polypropylene fumarate, poly(ethylglutamate-co-glutamic acid), poly(tert-butyloxy-carbonylmethylglutamate), polycaprolactone (PCL), polycaprolactone co-butylacrylate,polyhydroxybutyrate (PHBT) and copolymers of polyhydroxybutyrate,poly(phosphazene), polyphosphate ester), poly(amino acid),polydepsipeptides, maleic anhydride copolymers, polyiminocarbonates,poly[(97.5% dimethyl-trimethylene carbonate)-co-(2.5% trimethylenecarbonate)], poly(orthoesters), tyrosine-derived polyarylates,tyrosine-derived polycarbonates, tyrosine-derived polyiminocarbonates,tyrosine-derived polyphosphonates, polyethylene oxide, polyethyleneglycol, polyalkylene oxides, hydroxypropylmethylcellulose,polysaccharides such as hyaluronic acid, chitosan and regeneratecellulose, and proteins such as gelatin and collagen, and mixtures andcopolymers thereof, among others as well as PEG derivatives or blends ofany of the foregoing.

In the methods of removing or preventing biofilm formation, the surfacecan be a hard (for example, glass, metal, wood, chrome, plastic, vinylor formica) or a soft surface (for example, cloth or upholstery). Themethods set forth herein can be used to remove or prevent biofilmformation in vitro, ex vivo or in vivo. The methods set forth herein canalso be used to remove or prevent biofilm formation on a medical deviceor a part thereof. For example, the methods set forth herein can be usedto remove or prevent biofilm formation on an implantable medical devicesuch as a cardiac rhythm management device (for example, a pacemaker, adefibrillator, an implantable cardioverter defibrillator (ICD) and acardiac resynchronization therapy defibrillator (CRT device), aneurostimulator, a pulse generator, a drug pump, an infusion device, aphysiological monitoring device (for example, a glucose sensor), contactlenses, a stent, a catheter, tubing or a breast implant. Mesh, bandages,and implantable devices, for example, can be coated with a compositionscomprising one or more of the compounds set forth herein. Further,organs can be treated with one or more of the compounds set forth hereinprior to transplantation in a subject. One or more of the compounds setforth herein can be used to inhibit biofilm formation by one or more ofStaphylococcus aureus, Pseudomonas aeuroginosa, Staphylococcusepidermidis, Escherichia coli or Acinetobacter baummanii.

As utilized herein, by preventing biofilm formation is meant a method ofprecluding, delaying, averting, obviating, forestalling, stopping, orhindering the onset, incidence, severity, or recurrence of biofilmformation. For example, the disclosed method is considered to beprevention if there is about a 10% reduction in onset, incidence,severity, or recurrence of biofilm formation on a surface when comparedto a control surface that did not receive a composition for preventingbiofilm formation. Thus, the reduction in onset, incidence, severity, orrecurrence of biofilm can be about a 10, 20, 30, 40, 50, 60, 70, 80, 90,100%, or any amount of reduction in between as compared to a controlsurface.

A biofilm can also exist or form in a biological subject, for example,on the teeth or gums of a subject. Therefore, one or more of thecompounds set forth herein can be in a toothpaste, mouth rinse, gel,foam, varnish, polish, floss, dental strip, or copolymer membrane inorder to remove or prevent biofilm formation on a dental surface.

Further provided herein is a method of identifying an antimicrobialagent comprising contacting a bacterial culture with a test agent andmeasuring adenylate kinase release in the supernatant of the bacterialculture, wherein an increase in adenylate kinase release as compared toa control indicates that the test compound is an antimicrobial agent.The control can be a bacterial culture that was not contacted with thetest compound. The bacterial culture can be a culture of any bacterialstrain, for example, a culture of any of the bacteria disclosed herein.The bacterial culture can also be small colony variant bacterial cultureor a biofilm associated bacterial culture. Examples of agents identifiedutilizing this method are provided in the Examples.

Disclosed are materials, compositions, and components that can be usedfor, can be used in conjunction with, can be used in preparation for, orare products of the disclosed methods and compositions. These and othermaterials are disclosed herein, and it is understood that whencombinations, subsets, interactions, groups, etc. of these materials aredisclosed that while specific reference of each various individual andcollective combinations and permutations of these compounds may not beexplicitly disclosed, each is specifically contemplated and describedherein. For example, if a method is disclosed and discussed and a numberof modifications that can be made to a number of molecules including inthe method are discussed, each and every combination and permutation ofthe method, and the modifications that are possible are specificallycontemplated unless specifically indicated to the contrary. Likewise,any subset or combination of these is also specifically contemplated anddisclosed. This concept applies to all aspects of this disclosureincluding, but not limited to, steps in methods using the disclosedcompositions. Thus, if there are a variety of additional steps that canbe performed, it is understood that each of these additional steps canbe performed with any specific method steps or combination of methodsteps of the disclosed methods, and that each such combination or subsetof combinations is specifically contemplated and should be considereddisclosed.

Publications cited herein and the material for which they are cited arehereby specifically incorporated by reference in their entireties.

The following examples are put forth so as to provide those of ordinaryskill in the art with a complete disclosure and description of how thecompounds and/or methods claimed herein are made and evaluated, and areintended to be purely exemplary of the invention and are not intended tolimit the scope of what the inventors regard as their invention exceptas and to the extent that they are included in the accompanying claims.

Example I Identification of Antimicrobial Compounds

Adenylate kinase (AK) is a ubiquitous intracellular enzyme that isreleased into the extracellular space upon cell lysis. As shown herein,AK release serves as a useful reporter of bactericidal agent activityand can be exploited for antimicrobial screening purposes. The AK assayexhibits improved sensitivity over that of growth-based assays and candetect agents that are active against bacteria in clinically relevantgrowth states that are difficult to screen using conventionalapproaches, such as small colony variants (SCV) and bacteria withinestablished biofilms. The usefulness of the AK assay was validated byscreening a library of off-patent drugs for agents that exhibitantimicrobial properties toward a variety of bacterial species,including Escherichia coli and all members of the “ESKAPE” pathogens(Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae,Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacterspecies). The assay detected antibiotics within the library that wereexpected to be active against the organism screened. Moreover, 38 drugselicited AK release. Examples include, the antihistamine, terfenadine,which was active against S. aureus planktonic, SCV population, andbiofilm-associated cells, to name a few. Tamoxifen, an estrogen receptorantagonist, was active toward E. faecium in vitro and also reduced E.faecium pathogenesis in a Galleria mellonella infection model. Thesedata demonstrate that the A K assay provides an attractive screeningapproach for identifying new antimicrobial agents. Further, drugsidentified using this screening approach, for example, terfenadine andtamoxifen, provide novel antimicrobial drug development scaffolds.

The ESKAPE pathogens frequently cause health care-associated bacterialinfections and can escape the effects of most currently availableantibiotics. The most successful and widely applied method to identifyagents with antibacterial activity has been whole-cell, bacterial growthassays. In this approach, libraries of small molecules or naturalproducts are screened for agents that limit bacterial growth. However,growth-based assays have limitations. For example, the growth orno-growth readout has a limited dynamic range. This is likely to beproblematic because growth assays lack the sensitivity required todetect antimicrobial molecules that are present in low concentrationswithin complex natural product extract libraries or compounds withlimited antimicrobial activity. While the latter would obviously notrepresent a molecule that could be directly translated to clinical use,these low-activity hits could provide structurally novel scaffoldssuitable for medicinal chemistry-based optimization. In addition,traditional growth-based assays are not readily amenable to screeningfor agents that target bacteria within certain clinically relevantbacterial growth states, such as established biofilms and small-colonyvariants.

To address these limitations, provided herein is a high-throughputscreen (HTS)-compatible whole-cell assay to detect agents that directlykill bacteria. The assay is based on the release of intracellularadenylate kinase (AK) into culture medium as a reporter of bacterialcell death. As shown herein, the AK assay exhibits improved sensitivityover that of conventional whole-cell growth assays and displaysspecificity for bactericidal agents. Further, the assay can be used toscreen for agents that kill small-colony-variant bacteria and bacteriawithin established biofilms.

To validate the A K assay as an HTS-compatible screening platform, thePrestwick library of off-patent drugs was screened against E. coli andeach of the ESKAPE pathogens. This library contains representativeexamples of nearly all classes of antibiotics, and the bactericidalagents within the library that were expected to be active against theorganism screened were identified. Additionally, agents with nopreviously reported antibiotic activity were identified. Traditional MICtesting confirmed the antimicrobial properties of many of thesemolecules, showing that they could be repurposed as antimicrobials orserve as lead molecules for antibiotic development. Consistent with thatprediction, it was shown that one of these compounds, tamoxifen, isactive against E. faecium in a Galleria mellonella model of infection.Further, it was shown that terfenadine is active against planktonic,small-colony variant, and biofilm-associated S. aureus. Taken together,these data demonstrate that the AK assay provides a general approach toscreening for new antimicrobial agents active against a variety ofpathogens during planktonic and other disease-associated growth states.

Bacterial Growth Conditions—

The bacterial species and strains used in these experiments are listedin Table 1. S. aureus strain UAMS-1112 (generous gift from M. Smeltzer,University of Arkansas Medical Center) is a stable small-colony variantof the common laboratory S. aureus strain 8325-4, which harbors a hemBdeletion. Unless otherwise noted, bacteria were grown for 16 h inMueller-Hinton (MH) (Becton, Dickinson, Franklin Lakes, N.J.) or brainheart infusion (BHI) (Becton. Dickinson) medium at 37° C. on a rotaryshaker at 225 rotations per min (rpm) and then used to inoculate (1:100)fresh medium and processed, as described below.

TABLE 1 Bacterial Strains Source or Species Strain Relevantresistance^(a) reference^(b) Enterococcus 824-05 Amp, Cf, Cp, Cl, Mp,Clinical faecium Tmp, Sul, Erm, Kan isolate Staphylococcus USA300-0114Amp, Cf, Cl, Sul, Kan 6 aureus UAMS-1 ND 7 UAMS-1112 Erm Univ. ofArkansas RN4220 Cl 8 Klebsiella cKP1 Amp, Cp, Sul, Erm, 9 pneumoniae VanAcinetobacter 98-37-09 Amp, Lz 10  baumannii Pseudomonas PA01 Amp, Kan,Lz, Sul, 11  aeruginosa Van Enterobacter PMD1001 Amp, Erm, Lz, Sul,Clinical cloacae Van isolate Escherichia 8295 Erm, Lz, Sul Clinical coliisolate ^(a)Abbreviations: ampicillin. Amp; colistin, Cl; ceftriaxone,Cf; ciprofloxacin, Cp; erythromycin, Erm: kanamycin, Kan; linezolid, Lz:meropenem, Mp; minocycline, Min; Sulfamethoxazole, Sul; vancomycin, Van;not determined, ND; Univ., university. ^(b)Clinical isolates wereobtained from the University of Rochester School of Medicine andDentistry.

Chemicals—

The Prestwick Chemical Library of molecules with known biologicalactivities was acquired from Prestwick Chemical (Illkirch, France).ToxiLight BioAssay kits were obtained from Lonza (Basel, Switzerland).Terfenadine, suloctidil, clomiphene citrate, ceftriaxone,sulfamethoxazole, erythromycin, kanamycin, ciprofloxacin, rifampin,ampicillin, minocycline, tamoxifen, and trimethoprim were purchased fromSigma-Aldrich (St. Louis, Mo.). Meropenem, linezolid, and vancomycinwere purchased from Thermo Fisher (Waltham, Mass.). Colistin waspurchased from APP Pharmaceuticals (Schaumburg, Ill.).

MIC Testing—

MIC testing was performed to determine the antibiotic susceptibilityprofile of selected bacterial strains according to Clinical andLaboratory Standards (CLSI) protocols (Hindler et al. Antimicrobialsusceptibility testing, section 5. Clinical microbiology procedureshandbook, Vol. 1 America Society of Microbiology, Washington, DC (2010).Briefly, colonies of each bacterial species were collected from MH agarplates and suspended in individual tubes of MH medium to an opticaldensity (600 nm) of 0.8. The resulting cultures were incubated at 37° C.in a rotary shaker at 225 rpm to exponential phase (˜1×10⁸ CFU ml⁻¹) andthen diluted in fresh MH medium to a cell density of ˜3×10⁷ CFU ml⁻¹.Ten microliters of the diluted cultures was added to 88 μl of MH mediumin individual wells of a 96-well, round-bottom plate (Corning, Inc.),and 2 μl of a stock solution of the indicated reference antibiotic ortest compound (0 to 256 μg ml⁻¹) was added to each well. The carriersolvent was either water or dimethyl sulfoxide (DMSO); final DMSOconcentrations were less than or equal to 2%. Plates were incubated at37° C. for 24 h, and the MIC was defined as the lowest concentration ofantibiotic in which there was no visible cell pellet in the wells.

Heat-Killed Bacterial AK Release Assays—

Overnight cultures of E. coli strain 8295 or S. aureus RN4220 were usedto inoculate (1:100 dilution) 25 ml of fresh M H medium and grown at 37°C. on a rotary shaker at 225 rpm to exponential phase (˜1×10⁸ CFU ml⁻¹).Cells were pelleted by centrifugation (2,000×g) and resuspended in 2.5ml of sterile water. One milliliter of the resulting suspension wasboiled for 3 min and filter sterilized (0.45-μm filter) to remove celldebris. The filtrate was serially diluted in sterile water, and 100 μlof each dilution was added to individual wells of a white-walled,96-well plate (Corning, Inc., Corning, N.Y.). To measure the AK activityin the supernatants at each dilution, 100 μl of ToxiLight AK reagent wasadded to each well, followed by incubation at room temperature for 30min, and luminescence was measured using a SpectraMax M5 plate reader(Molecular Devices, Sunnyvale, Calif.). Cells were serially diluted andplated to enumerate CFU (CFU ml⁻¹) before and after boiling to correlatecell lysis with viability.

AK Assay 96-Well Format—

Overnight cultures of each bacterial species were used to inoculate(1:100 dilution) 25 ml of fresh MH medium and grown at 37° C. on arotary shaker at 225 rpm to exponential phase (˜1×10⁸ CFU ml⁻¹).Ninety-eight microliters of MH medium. 2 μl of the indicated antibiotic,and 5×10⁶ bacteria were added to individual wells of a white-walled,96-well microtiter plate. Well components were mixed by pipetting andincubated at 37° C. for 3 h. The plate was equilibrated to roomtemperature for 30 min. Next, 100 μl of ToxiLight AK reagent was addedto each well and incubated at room temperature for 30 min, andluminescence was measured using a SpectraMax M5 plate reader.

AK Assay 384-Well Format and High-Throughput Screening—

Overnight cultures of each bacterial species were used to inoculate(1:100 dilution) 25 ml of fresh medium and grown at 37° C. on a rotaryshaker at 225 rpm to exponential phase (˜1×10⁸ CFU ml⁻¹). In awhite-walled, 384-well plate, 24 μl of MH medium, 0.3 μl (50 μM) ofantibiotic or compound, and 5×10⁶ bacteria were added to individualwells and incubated at 37° C. for 3 h. The plates were equilibrated atroom temperature for 1 h. Twenty-five microliters of ToxiLight AKreagent was then added to each well, followed by incubation at roomtemperature for 30 min, and luminescence was measured using a SpectraMaxM5 plate reader.

AK Assay of Established Bloflims—

Biofilms were grown as previously described (Beenken et al. Infect.Immun. 71: 4206-4211 (2003); Musken et al. Nat. Protoc. 5: 1460-1459(2010); Tomaras et al. Microbiology 149: 3473-3483 (2003)). Briefly, P.aeruginosa and A. baumannii were cultured overnight in Luria-Bertanimedium and then used to seed 96-well, flat-bottom plates. Plates wereincubated at 37° C. in a humidified incubator for 48 h to allow theformation of static biofilms. Nonadherent cells were removed byaspiration and washing with sterile phosphate-buffered saline (PBS).Fresh LB medium supplemented with 0, 1×, 10×, or 100×MIC of antibioticwas added to each well and incubated overnight at 37° C. Followingtreatment, 100 μl of each biofilm supernatant was transferred to96-well, white-walled plates, 100 μl of ToxiLight AK reagent was addedto each well, mixtures were incubated for 30 min at room temperature,and luminescence was measured using a SpectraMax M5 plate reader.Biofilm-associated bacteria were enumerated by resuspending each biofilmin fresh PBS and plating. For S. aureus UAMS-1 biofilms, 96-well,flat-bottom plates were first coated with 100 μl of 20% human plasma incarbonate buffer overnight at 4° C. Following coating, the plasmasolution was removed and cells were inoculated in each well 1:200 in 100μl of tryptic soy broth supplemented with 3% glucose and 0.5% NaCl.Biofilms were cultured for 48 h in a humidified incubator at 37° C.Established S. aureus biofilms were washed once with PBS and thentreated with a 100 μl of ToxiLight lysis buffer for 3 h, after which theamount of AK released into supernatants was measured, as describedabove.

Small-Colony Variant AK Assays—

Thirty-six-hour cultures of S. aureus strain UAMS-1112 were used toinoculate (1:100 dilution) 100 ml of fresh MH medium and grown at 37° C.on a rotary shaker at 225 rpm to an optical density (600 nm) of 0.1 to0.2, corresponding to ˜1×10⁶ CFU ml⁻¹. Cells were pelleted bycentrifugation and resuspended in 2 ml of fresh MH medium. Ninety-eightmicroliters of MH medium containing 5×10⁶ bacteria and 2 μl of theindicated antibiotic were added to individual wells of a white-walled,96-well microtiter plate. Well components were mixed by pipetting andincubated at 37° C. for 3 h. The plate was equilibrated to roomtemperature for 30 min. Next, 100 μl of ToxiLight AK reagent was addedto each well, followed by incubation at room temperature for 30 min, andluminescence was measured using a SpectraMax M5 plate reader.

Galleria mellonella Model of S. aureus Infection—

A Galleria mellonella model of infection was used to measure theputative antimicrobial properties of tamoxifen against E. faecium andterfenadine against S. aureus. To do so, overnight cultures of E.faecium strain 824-05 or S. aureus strain USA300-0114 were used toinoculate (1:100 dilution) 25 ml of fresh MH medium and grown at 37° C.on a rotary shaker at 225 rpm to exponential phase (˜1×10⁸ CFU ml⁻¹).Cultures were pelleted by centrifugation (2,000×g), washed with sterilePBS, and resuspended at ˜1×10⁹ CFU ml⁻¹ in fresh PBS. Galleriamellonella larvae (Vanderhorst Wholesale. Inc.; St. Marys, Ohio)weighing 200 to 300 mg were inoculated with 5 μl of E. faecium or S.aureus (5×10⁶ CFU) into the last left proleg using a 10-μl Hamiltonsyringe. Worms were then mock treated with either DMSO (negativecontrol), vancomycin (20 mg kg⁻¹: positive control) at 2 h and 24 hpostinoculation. For E. faecium studies, groups were also treated withthe test compound tamoxifen at 80, 160, or 320 mg kg⁻¹, whereas groupswere treated with the test compound terfenadine (80, 160, or 320 mgkg⁻¹) for S. aureus studies. Treatments were administered in the samemanner as infection, except that each injection was in the next leftproleg moving toward the head of the worm. Larvae were housed in petridishes in the dark at 37° C. and monitored for viability at theconclusion of the study (48 h postinoculation); worms were considereddead if they did not respond to physical stimuli. In addition to mock orcompound treatment of infected larvae, studies included two additionalnoninfected negative-control groups: one group that did not receiveinjections and one group that was injected with PBS to control for theimpact of physical trauma. All experimental groups contained 15 worms,and each experiment was repeated three times.

Rationale for Adenylate Kinase as a Reporter of Bacterial Cell Lysis—

Adenylate kinase (AK) is a ubiquitous intracellular enzyme thatcatalyzes the conversion of 2 ADP⇄ATP+AMP and is released into theextracellular space upon cell lysis. The premise of the assay is thatagents which disrupt cellular integrity, either directly through damageof the membrane/cell wall or indirectly following the death of the cell,will induce release of AK into the culture medium. Extracellular AK issubsequently detected by the addition of commercially availableToxiLight AK assay reporter cocktail (Lonza, Basel, Switzerland), whichgenerates a luminescent signal by utilizing AK-generated ATP in thestandard luciferase catalyzed reaction (see FIG. 1). As shown herein, anAK assay was developed as a high-throughput screening platform forantibacterial drug discovery.

The AK Assay Provides a Sensitive Measure of Bacterial Lysis—

As an initial test of AK release as a reporter of bacterial cell deathfor Gram-positive and Gram-negative organisms, the sensitivity withwhich the assay measures AK in the culture supernatants of heat-killedE. coli and S. aureus was determined. Each bacterial species was grownto exponential phase, harvested, and resuspended at 1×10⁹ CFU per ml inMueller-Hinton (MH) medium. Bacterial suspensions were heat killed, anda 10-fold dilution series of supernatants was prepared; an aliquot ofeach heat-killed sample was plated to ensure ≧99% bacterial death. TheAK activity of the dilution series was measured and compared to the AKactivity of mock-treated (viable) bacteria by Student's t test. Incomparison to untreated cells, a statistically significant increase inAK activity was detected at dilutions containing 1.7×10³ or moreheat-killed E. coli supernatants, compared to results for live bacteria(see FIG. 2A). AK activity increased ˜433-fold and ˜1,574-fold incultures with 1×10⁷ and 1×10⁸ heat-killed E. coli supernatants,respectively, compared to results for mock-treated bacteria. Acomparison of AK released from heat-killed S. aureus to that ofmock-treated cells revealed a statistically significant difference in AKactivity was detected at dilutions containing 1.8×10⁴ or moreheat-killed S. aureus supernatants, with a maximum 122-fold increase inAK activity observed for 1×10⁸ lysed cells. Taken together, theseresults indicate that the AK assay reproducibly detects AK releasefollowing bacterial cell lysis and that the assay is extremelysensitive, allowing the identification of agents that cause lysis in0.0001% or 0.001% of the starting inoculum of E. coli or S. aureuscells, respectively. Furthermore, the AK assay exhibited a dynamic rangeof nearly 3 orders of magnitude and an excellent signal-to-noise ratio.

The AK Assay Detects Bactericidal Molecules that are Active Against theESKAPE Pathogens—

Next, ability of the AK assay to detect the activities of bactericidalantibiotics toward E. coli and each of the ESKAPE pathogens wasexamined. To do so, the MICs of six classes of bactericidal antibiotics(penicillin, cephalosporin, quinolone, glycopeptide, carbapenem, andpolymyxin) were determined for each organism (see Table 2). An AK assaywas then performed at 0, 0.125×, 0.25×, 0.5×, 1×, and 2×MIC of eachantibiotic. As discussed below, AK release was detected at antibioticconcentrations below the MIC for most antibiotics tested, showing thatthe AK assay is more sensitive than growth-based assays for detectingbactericidal agents. As a representative example, FIG. 2B presents theAK assay measured polymyxin (colistin)-mediated killing of A. baumanniistrain 98-37-09. The MIC of colistin for this strain is 4 μg ml⁻¹,whereas AK release was robustly detected at 0.25×, 0.5×, and 1×MICvalue.

TABLE 2 MIC and AK measures of bacterial species and antibioticcombinations^(a) Value for agent Bactericidal Bacteriostatic Strain andTreatment Ceftri- Cipro- Vanco- Sulfame- Mino- Erythro- Line- parameter(×MIC) Ampicillin axone floxacin mycin Meropenem Colistin thoxazolecycline mycin Kanamycin zolid Enterococcus faecium 824-05 MIC >256 >256256 2 >256 >256 >256 1 >256 >256 16 Fold increase 0.5 ND ND ND 1 ND NDND 3 ND ND 2 in AK signal 1.0 ND ND ND 2 ND ND ND 4 ND ND 2Staphylococcus aureus USA300-0114 MIC >256 >256 2 1 4 >256 >256 164 >256 2 Fold increase 0.5 ND ND 3 11 13 ND ND 1 1 ND 1 in AK signal1.0 ND ND 5 3 17 ND ND 1 1 ND 1 RN4220 MIC 2 2 2 2 2 >256 2 8 2 16 2Fold increase 0.5 4 10 8 3 11 ND 2 1 1 1 1 in AK signal 1.0 3 12 10 6 11ND 3 1 1 1 1 Klebsiella pneumoniae cKP1 MIC >256 2 >256 >256 2 8 >25664 >256 2 >256 Fold increase 0.5 ND 50 ND ND 128 34 ND 1 ND 1 ND in AKsignal 1.0 ND 55 ND ND 220 33 ND 1 ND 1 ND Acinetobacter baumannii98-37-09 MIC 256 8 0.0625 64 2 4 8 1 16 2 256 Fold increase 0.5 ND 5 2 279 17 6 23 3 3 ND in AK signal 1.0 ND 21 2 4 134 49 5 14 12 14 NDPseudomonas aeruginosa PA01 MIC >256 4 2 >256 2 4 256 32 32 256 >256Fold increase 0.5 ND 2 27 ND 45 5 ND 1 3 ND ND in AK signal 1.0 ND 14182 ND 39 12 ND 6 1 ND ND Enterobacter cloacae PMD1001 MIC >256 2<1 >256 2 2 >256 4 256 2 >256 Fold increase 0.5 ND 20 6 ND 99 10 ND 4 ND1 ND in AK signal 1.0 ND 17 7 ND 296 32 ND 9 ND 1 ND Escherichia coli8295 MIC 128 2 0.125 16 2 8 >256 16 256 32 >256 Fold increase in 0.5 5051 17 1 60 15 ND 4 ND 55 ND AK signal 1.0 74 76 39 2 85 22 ND 38 ND 70ND % increase 0.5 100 83.3 83.3 40 100 100 50 50 50 40 0 expected 1.0100 100 83.3 60 100 100 100 62.5 25 40 0 ^(a)Shading indicatessignificant increase in AK signal (greater than or equal to threefoldover vehicle-treated cells).

The MIC measures for each ESKAPE pathogen and each organism'scorresponding fold increase in AK signal following treatment with 0.5×and 1.0× the MIC are provided in Table 2 (bactericidal agents). For fourof the six bactericidal antibiotics tested, the AK assay proved to besuperior to growth-based assays with respect to detecting the killingproperties of bactericidal agents at sub-MIC values. More specifically,100% of the organisms that were determined to be susceptible to the cellwall-active antibiotics ampicillin, meropenem, and colistin exhibited asignificant increase in AK signal (≧3-fold over that for vehicle-treatedcells) at both 0.5× and 1.0× their MIC's. Eighty-three percent and 100%of ceftriaxone-susceptible species exhibited increased AK signal at 0.5×and 1× their MICs, respectively. Five of six (83%)ciprofloxacin-susceptible organisms exhibited AK signal at both 0.5× and1.0×MIC. Interestingly, the cell wall-targeted glycopeptide, vancomycin,caused significant AK release at 0.5× and 1.0× their MICs in only 40%and 60% of susceptible species.

To evaluate the specificity of the assay for detecting bactericidalagents, measured AK release by each of the ESKAPE pathogens followingexposure to five classes of bacteriostatic agents (sulfonamide,tetracycline, macrolide, aminoglycoside, and oxazolidine) was alsomeasured. To do this, the MIC value of each bacteriostaticantibiotic/organism pair was measured by a conventional growth-basedapproach. AK release by each bacterial species following treatment with0.5× or 1.0×MIC of each bacteriostatic agent was subsequently measured.As shown in FIG. 3, S. aureus strain RN4220 was susceptible to allagents tested but bacteriostatic agents generated very low AK release,whereas bactericidal antibiotics generated robust AK detection. In mostinstances, a similar trend was observed for the other ESKAPE pathogens,indicating that the assay enriches for the identification ofbactericidal agents, which are arguably the most valuable antibioticsbecause they can be used to treat patients with immunological defects orrapidly lethal infections (Table 2, bacteriostatic agents). Twoexceptions were noted. Surprisingly, A. baumannii generated significantAK signal in response to all the bacteriostatic agents evaluated,showing that AK release may be a feature of a more general stressresponse in this organism. Additionally, the bacteriostatic agentminocycline generated signal by five of the eight (63%) organismstested.

Taken together, these results show that the AK assay provides a viablescreening approach for identifying bactericidal agents at sub-MICs thatcould otherwise be missed by growth-based assays. Further, because theAK assay relies on bacterial killing as opposed to growth changes togenerate its readout, it was hypothesized that it would provide a formatto develop screens that are not readily available by conventional,growth-based approaches.

Use of the AK Assay to Identify Agents with Antimicrobial ActivitiesAgainst Established Biofilms and Small Colony Variants—

Established bacterial biofilms represent a particularly problematicdisease state, in part because biofilm-associated bacteria arerecalcitrant to conventional antibiotic therapy. Thus, they have been afocus of antibiotic development. As a result, a number of approaches toscreening for molecules with activity toward bacterial biofilms havebeen developed recently (Benoit et al. Environ. Microb. 76: 4136-4142;Perez et al. Lett. Appl. Microb. 51:331-337 (2010)). Although each hasits advantages, they also have a number of limitations, includingreproducibility, reliance on specialized equipment, or low throughput.It was hypothesized that the AK assay would provide a solution to someof these problems because it is rapid, sensitive, and simple to performand it detects bactericidal molecules, the type of antibiotics requiredto treat established biofilms.

To determine whether the AK assay could detect agents with activityagainst established biofilms, S. aureus strain UAMS-1, A. baumanniistrain 98-37-09, and P. aeruginosa strain PAO1 static biofilms wereformed in 96-well flat-bottom plates. Forty-eight hours postinoculation,one well corresponding to each organism was stained with crystal violetto verify that biofilm formation had occurred, whereas the remainingwells were treated with 10× the MIC value with either colistin (A.baumannii biofilms) or ciprofloxacin (P. aeruginosa, and S. aureusbiofilms). Following overnight antibiotic treatment, biofilm-associatedbacteria were enumerated by plating, and the corresponding supernatantswere analyzed by the AK assay. Plating verified that 10×MIC antibiotictreatment resulted in a significant reduction in biofilm-associated P.aeruginosa (a 3.1-log decrease), S. aureus (a 0.7-log decrease), and A.baumannii (a 1.8-log decrease) compared to findings for untreatedbiofilms. As shown in FIG. 4A, corresponding AK measures indicated thatthe assay robustly detects the mild effects of these antibiotics on eachbacterial species tested, showing that it represents a promisingapproach to identify agents that exhibit bactericidal activity towardestablished bacterial biofilms.

Another context in which the AK assay can be particularly valuable is inthe identification of molecules that exhibit bactericidal activitytoward bacterial small-colony variants (SCV). SCV are slow-growingpopulations of bacterial species that have been hypothesized to causelatent or recurrent infections and are tolerant of standard antibiotictreatment regimens. Based on the aberrant SCV growth characteristics,typical growth-based HTS assays would be difficult to employ.Accordingly, the ability of the AK assay to identify agents that kill S.aureus SCV strain UAMS-1112 was tested. To do so, 1×10⁶ UAMS-1112 cellswere treated with 1× or 10×MIC ciprofloxacin, meropenem, or vancomycinfor 3 h. Following treatment, suspensions were plated to measure theantimicrobial properties of each antibiotic, and the AK assay wasperformed to measure adenylate kinase release. Plating revealed thatciprofloxacin and vancomycin had no effect on SCV viability at anyconcentration tested and exhibited no change in AK release in comparisonto results for mock-treated cells (FIG. 4B), supporting the observationthat S. aureus small-colony variants are recalcitrant to antibiotictreatment. Meropenem treatment resulted in a 0.5-log decrease in SCVviability, which corresponded to a 2.5-fold increase in AK signalcompared to that for mock-treated cells (FIG. 4B). Taken together, theseresults show that the AK assay provides the sensitivity needed to detectthe slight antimicrobial effects of antibiotics, such as meropenem,toward S. aureus SCV.

Validation of AK as an HTS-Compatible Assay of Antibacterial Activity—

To test the viability of the AK assay as a general tool in HTS-basedantibacterial small-molecule discovery, optimized assay parameters,including inoculum, drug incubation time, and AK reaction time, forscreening planktonic ESKAPE species as well as E. coli in a 384-wellformat were determined. Based on these experiments, a standardizedassay, as described above, was developed. As part of this optimizationprocess, control assays in a 384-well format were performed to measurethe signal to noise and reproducibility of the assay in an I-ITS manner.For this, plates were seeded with E. coli or an ESKAPE pathogen.Alternating columns of the plate were then mixed with either 2% DMSO(negative control) or a bactericidal antibiotic (positive control), andAK signal was detected; a representative result for DMSO- andcolistin-treated K. pneumoniae is shown in FIG. 5A. Comparisons of thevariance in signal between positive- and negative-control measuresindicated that the AK assay provides Z′-factor scores between 0.59 and0.82 (depending on the specific organism), indicating that the assay issufficiently robust for HTS.

AK-Based Screening of Library of Off-Patent Drugs and BiologicallyActive Molecules—

To further validate the AK assay protocol, E. coli and each of theESKAPE pathogens was screened against the Prestwick library ofFDA-approved drugs and biologically active molecules (1,120 compounds,total; 50 μM final drug concentration). The Prestwick library containsrepresentatives of nearly all classes of antibiotics currently inclinical use, making it an ideal library for testing the ability of theAK assay to detect bactericidal agents in a high-throughput screeningformat. Accordingly, the library was screened for antimicrobial agentsthat were active against planktonic E. coli and each of the ESKAPEpathogens using the AK assay; the cutoff for positive-scoring moleculeswas set at a 3-fold increase in extracellular AK activity, correspondingto the detection limit of statistically significant increases in AKactivity for planktonic bacteria. The hit rates for the differentorganisms ranged from 1.4% to 4.8%; a representative example of rawscreening data for Klebsiella pneumoniae is shown in FIG. 5B. Screeningresults for all ESKAPE pathogens are summarized in Table 3, whereasresults for all compounds within the Prestwick library are provided inTable 4.

TABLE 3 Preswick library screening results No. of active agents forspecies Compound E. coli E. faecium S. aureus ^(a) K. pneumoniae A.baumannii ^(a) P. aeruginosa E. cloacae Bactericidal 45 4 25 19 17 21 16Bacteriostatic 2 1 14 0 7 0 1 Detergent 2 2 2 2 2 1 0 Other 5 9 10 4 181 12 Total 54 16 51 25 44 23 29 ^(a)Tetracyclines were identified.

TABLE 4 AK-based Prestwick Library Screening Results. E. coli E. faeciumS. aureus K. pneumoniae A. baumannii P. aeruginosa E. cloacae Chemicalname Mechanism Therapeutic group 0.4 0.8 0.6 0.3 0.3 0.3 0.3Azaguanine-8 Purine antimetabolite Antineoplastic 0.4 0.8 0.4 0.4 0.40.3 0.3 Metronidazole Bacterial DNA damage Antibacterial 0.4 0.7 1.4 0.40.4 0.3 0.3 Allantoin Precipitate proteins Vulnerary 0.5 0.8 0.4 0.4 0.30.3 0.3 Cotinine (−) Antidepressant 0.4 0.9 0.4 0.4 0.4 0.3 0.3Acetazolamide Carbonic anhydrase inhibitor Diuretic 0.4 0.8 0.4 0.5 0.40.4 0.3 Edrophonium chloride Cholinergic Myasthenia Gravis test 0.5 0.80.4 0.4 0.4 0.3 0.3 Metformin hydrochloride — Antidiabetic 0.5 0.8 0.40.4 0.4 0.3 0.3 Moroxidine hydrochloride — Antiviral 0.5 0.8 0.5 0.5 0.40.4 0.3 Atracurium besylate Neuromuscular blocking agent Curarizingagent 0.5 0.8 0.4 0.6 0.4 0.3 0.3 Baclofen (R,S) GABAb agonistAntispasmodic 0.5 0.9 0.4 0.5 0.4 0.4 0.4 Isoflupredone acetateAnti-inflammatory 0.5 0.9 0.5 0.5 0.4 0.4 0.3 Acyclovir ADN polymeraseinhibitor Antiviral 0.5 0.8 0.4 0.4 0.4 0.3 0.4 Amiloride hydrochloridedihydrate Antialdosteron Diuretic 0.5 0.9 0.5 0.4 0.4 0.4 0.4 DiazoxideActivator of ATP-dependent K+ channels Antihypertensor 0.5 0.9 0.5 0.60.5 0.4 0.4 Amprolium hydrochloride Thiamine transport inhibitorCoccidiostatic 0.5 0.9 0.4 0.5 0.4 0.4 0.4 Amidopyrine ACTH secretorAntipyretic 0.5 1.1 0.5 0.6 0.5 0.5 0.4 Hydrochlorothiazide Na+ Cl−transport inhibitor Diuretic 0.4 2.8 0.1 0.4 0.5 0.4 0.5 Ursolic acid —Diuretic 0.5 1.0 0.5 0.6 0.5 0.5 0.4 Sulfaguanidine Inhibitor of folicacid synthesis Antibacterial 0.5 1.0 0.5 0.6 0.5 0.4 0.4 Pindolol Betaadrenergic antagonist Antiarrhythmic 0.6 1.0 0.7 0.6 0.6 0.6 0.6Isoniazid — Antibacterial 0.6 0.9 0.7 0.6 0.5 0.7 0.7 Mexiletinehydrochloride Na+ channel blocker Antiarrhythmic 0.6 1.0 0.7 0.5 0.6 0.70.6 Pentylenetetrazole GABA antagonist CNS stimulant 0.5 0.9 0.6 0.6 0.50.6 0.6 Flavoxate hydrochloride Phosphodiesterase inhibitorAntispasmodic 0.6 0.9 0.5 0.5 0.6 0.6 0.6 Chlorzoxazone — Musclerelaxant 0.5 0.9 0.4 0.5 0.5 0.5 0.6 Bufexamac — Antiinflammatory 0.60.9 0.5 0.5 0.6 0.6 0.6 Ornidazole Bacterial DNA damage Antibacterial0.5 0.9 0.5 0.6 0.6 0.6 0.5 Glutethimide, para-amino Aromatase inhibitorAntineoplasic 0.6 0.9 0.5 0.6 0.5 0.5 0.6 Ethosuximide Ca++ channelinhibitor Anticonvulsant voltage dependant 0.7 0.9 0.5 0.5 0.5 0.5 0.5Dropropizine (R,S) — Antitussive 0.6 1.2 0.6 0.5 0.5 0.5 0.6 Mafenidehydrochloride Inhibitor of folic acid Antibacterial biosynthesis 0.5 0.90.5 0.5 0.5 0.5 0.6 Pinacidil K+ channel Ca++ dependant Vasodilatatoractivator 0.2 0.3 0.2 0.2 0.2 0.2 0.5 Riluzole hydrochloride Glutamateantagonist Neuroprotective 0.3 0.5 0.2 0.3 0.3 0.3 0.4 Albendazole 0.41.0 0.4 0.8 0.5 0.4 0.6 Nitrofurantoin Bacterial DNA damage Urinaryantiseptic 0.5 0.9 0.5 0.5 0.6 0.5 0.6 Clonidine hydrochloride Alpha2agonist Antihypertensor 0.0 0.3 0.2 0.0 0.1 0.0 0.3 Hydralazinehydrochloride Adrenergic antagonist Antihypertensor 0.5 1.0 0.5 0.5 0.50.5 0.5 Bupropion hydrochloride — Antidepressant 0.5 0.8 0.5 0.6 0.5 0.50.5 Phenelzine sulfate MAO inhibitor Antidepressant 0.6 0.9 0.6 0.5 0.70.5 0.4 Alprenolol hydrochloride Beta1 antagonist Antihypertensor 0.60.9 0.8 0.5 0.8 0.6 0.7 Meticrane Na+ channel blocker Antihypertensor0.6 0.9 0.7 0.6 0.7 0.7 0.7 Khellin — Phototherapeutic agent 0.7 0.8 0.70.6 0.7 0.6 0.7 Benzonatate Antitussive 0.7 0.9 0.8 0.5 0.7 0.6 0.8Zimelidine dihydrochloride 5-HT uptake inhibitor Antidepressantmonohydrate 0.7 0.9 0.5 0.5 0.7 0.5 0.7 Hydroflumethiazide Na+ Cl−transport inhibitor Antihypertensor 0.6 0.9 0.7 0.6 0.6 0.6 0.9Azacyclonol H1 antagonist Anxiolytic 0.6 0.9 0.7 0.5 2.2 0.5 0.7Sulfacetamide sodic hydrate Antipsoriasic 0.6 0.8 0.7 0.5 0.7 0.6 0.8Azathioprine Antimetabolite Immunosuppressant 0.8 0.9 0.7 0.5 0.6 0.50.7 Heptaminol hydrochloride — Antihypotensive 0.7 0.9 0.5 0.6 1.4 0.60.7 Lynestrenol Antioestrogen Progestogen 0.7 0.9 0.6 0.6 1.0 0.6 0.7Sulfathiazole Inhibitor of folic acid Antibacterial synthesis 0.7 1.00.5 0.4 0.8 0.6 0.7 Guanabenz acetate Alpha agonist Antihypertensor 0.60.8 0.5 0.6 0.7 0.5 0.8 Levodopa Tyrosine aminotransferaseAntiparkinsonian inhibitor 0.9 1.0 1.1 0.6 1.0 0.6 0.7 DisulfiramDopamine beta-hydroxylase Alcohol deterrent inhibitor 0.7 1.0 0.7 0.60.7 0.6 0.8 Idoxuridine Nucleic acid synthesis Antiviral inhibitors 0.61.0 0.7 0.6 0.7 0.6 0.8 Acetylsalicylsalicylic acid Cyclooxygenaseinhibitor Antiinflammatory 0.7 1.0 0.7 0.6 0.8 0.6 0.8 CaptoprilAngiotensive converting Antihypertensor enzyme inhibitor 0.7 0.9 0.7 0.60.8 0.8 0.7 Mianserine hydrochloride 5-HT antagonist Antidepressant 0.61.0 0.7 0.6 0.8 0.6 0.7 Minoxidil K+ channel activator Antihypertensor0.6 0.9 0.6 0.5 0.6 0.6 0.7 Nocodazole Microtubule poison Antineoplastic0.8 1.1 0.7 0.7 0.8 0.9 0.9 Tranexamic acid Plasminogen inhibitorHemostatic 0.9 1.0 0.8 0.7 0.9 0.9 0.9 Chlorothiazide Na+ Cl− transportinhibitor Antihypertensor 0.8 1.0 0.8 0.6 0.8 0.7 0.9 EtofyllinePhosphodiesterase inhibitor Cardiac analeptic 0.8 1.0 0.7 0.7 0.7 0.80.8 Diphenidol hydrochloride — Antivertigo 0.8 1.0 0.8 0.6 0.9 0.8 0.9Tranylcypromine hydrochloride MAO inhibitor Antidepressant 0.7 1.0 0.80.7 0.7 0.8 0.9 Norethindrone — Progestogen 0.9 0.9 0.7 0.7 0.9 0.8 0.9Alverine citrate salt Anticholinergic Spasmolytic 0.9 0.9 1.0 0.7 0.90.8 0.9 Nortriptyline hydrochloride Norepinephrine uptake Antidepressantinhibitor 0.7 1.0 0.7 0.6 0.7 0.8 0.8 Aceclofenac Cyclooxygenaseinhibitor Antiinflammatory 0.6 0.9 0.7 0.6 0.7 0.7 0.8 Niflumic acidProstaglandine synthesis Analgesic inhibitor 0.8 1.0 1.0 0.7 0.7 0.7 0.8Iproniazide phosphate Monoamine oxydase Antidepressant inhibitor (nonselective) 0.7 1.0 0.6 0.6 0.7 0.8 0.9 Isotretinoin — Cystic acne 0.71.0 0.7 0.6 1.3 0.7 0.7 Sulfamethoxazole 0.7 1.0 0.8 0.6 0.8 0.7 0.9Retinoic acid — Keratolytic 0.7 1.0 0.8 0.7 0.8 0.6 0.8 Mephenesin —Muscle relaxant 0.7 1.0 0.8 0.6 0.8 0.7 0.8 Antazoline hydrochloride H1antagonist Antihistaminic 0.8 1.0 0.7 0.6 0.8 0.7 0.8 Phenforminhydrochloride Neoglucogenese inhibitor Antidiabetic 0.7 1.3 1.4 0.6 0.80.7 0.8 Ethacrynic acid Na+ Cl− uptake inhibitor Diuretic 0.7 1.0 0.40.6 0.7 0.8 0.7 Flutamide Androgenic receptor Anticancer antagonist 0.91.0 0.8 0.6 0.7 0.6 0.7 Praziquantel Modulates cell membraneAnthelmintic permeability 0.8 0.8 0.8 0.7 1.4 0.8 0.8 SulfaphenazoleInhibitor of folic acid Antibacterial synthesis 0.8 0.8 0.9 0.8 0.8 0.60.9 R(−) Apomorphine hydrochloride D1 agonist Emetic hemihydrate 0.9 0.80.6 0.7 0.8 0.7 1.0 Panthenol (D) — Vitamin 1.0 0.9 0.7 0.8 0.8 0.9 0.9Amoxapine Dopamine-reuptake Antidepressant inhibitor 1.0 0.8 0.7 0.7 1.20.8 0.9 Sulfadiazine Inhibitor of folic acid Antibacterial synthesis 0.90.9 0.6 0.6 0.9 0.9 0.9 Cyproheptadine hydrochloride 5-HT antagonistAntipruritic 0.9 0.9 0.7 0.7 0.7 0.8 1.0 Norethynodrel — Progestogen 0.90.8 0.5 0.8 0.8 0.8 1.0 Famotidine H2 histaminic antagonistAntiulcerative 0.9 0.9 8.7 0.4 0.9 0.8 1.4 Thiamphenicol Ribosomalprotein synthesis Antibacterial inhibitor 0.7 0.9 0.6 0.6 0.9 0.8 0.9Danazol Estrogen antagonist Antigonadotropin 0.9 0.9 0.8 0.7 0.8 0.8 1.0Cimetidine H2 antagonist Antiulcer 0.9 0.9 0.6 0.8 0.8 0.7 0.9Nicorandil Antihypertensor 0.8 0.9 0.7 0.6 0.8 0.8 1.0 Doxylaminesuccinate H1 antagonist Antihistaminic 0.9 1.0 0.8 0.7 1.0 0.8 1.1Tomatine Antifungal 0.9 0.9 0.6 0.7 0.8 0.7 1.1 Ethambutoldihydrochloride Chelating agent Antibacterial 0.9 0.9 0.7 0.8 0.8 0.81.0 Nomifensine maleate Dopamine uptake inhibitor Antidepressant 0.9 1.00.7 0.7 1.0 0.8 1.0 Antipyrine — Analgesic 0.9 0.9 0.7 0.7 0.8 0.9 1.0Dizocilpine maleate Probe for NMDA receptors — 0.9 1.0 0.7 0.7 0.8 0.81.0 Antipyrine, 4-hydroxy — Antipyrine metabolite 0.9 1.0 0.8 0.7 0.80.9 0.9 Acenocoumarol Vitamin K antagonist Anticoagulant 0.9 1.0 0.8 0.70.9 1.0 1.0 Ampyrone — Analgesic 1.0 0.9 0.8 0.8 0.9 1.0 1.1 Ethisterone— Progestogen 1.0 1.0 0.9 0.7 1.1 1.0 1.0 Levamisole hydrochlorideAlkaline phosphatase Immunomodulator inhibitor 1.0 1.0 0.6 0.7 0.9 1.01.0 Triprolidine hydrochloride H1 antagonist Antihistaminic 1.0 0.9 0.80.8 0.9 1.0 0.9 Pargyline hydrochloride Monoamine oxidaseAntihypertensor inhibitor 1.1 1.0 0.9 0.8 0.9 1.1 1.0 Doxepinhydrochloride Adrenaline uptake inhibitor Anticonvulsant 0.9 0.9 0.8 0.60.9 0.9 1.0 Methocarbamol — Muscle relaxant 1.0 1.0 0.9 0.7 0.9 1.0 1.1Dyclonine hydrochloride Na+ channel blocker Local anesthesic 15.9 1.00.6 2.6 5.4 9.1 18.4 Aztreonam Bacterial transpeptidase Antibacterialinhibitor 1.0 1.0 0.8 0.7 0.8 1.0 1.2 Dimenhydrinate H1 antogonistAntihistaminic 1.0 1.0 5.3 0.7 1.3 1.0 1.0 Cloxacillin sodium saltBacterial transpeptidase Antibacterial inhibitor 0.9 0.9 2.7 0.6 0.9 0.91.0 Disopyramide Na+ channel blocker Antiarrhythmic 1.0 1.0 0.8 0.8 0.80.7 1.0 Catharanthine 1.0 1.3 2.0 1.0 3.3 1.0 1.0 Clotrimazole Specificinhibitor of Ca2+ Antibacterial activated K+ channels 0.8 0.9 0.7 0.71.0 1.0 1.1 Pentolinium bitartrate Glanglionic blocking agentAntihypertensor 0.9 1.0 0.8 0.8 0.9 1.0 1.0 VinpocetinePhosphodiesterase inhibitor Nootropic drug 0.9 0.9 0.7 0.8 0.8 0.9 1.1Aminopurine, 6-benzyl Inhibitor of respiratory kinase in plants 1.1 1.10.8 0.7 1.0 0.9 1.0 Clomipramine hydrochloride Noradrenaline reuptakeAntidepressant inhibitor 0.8 1.0 0.7 0.8 0.9 0.9 1.0 Tolbutamide K+channel ATP dependant Hypoglycemic inhibitor 1.0 1.1 0.8 0.8 1.5 1.1 0.9Fendiline hydrochloride Ca++ channel activator Antianginal 0.7 0.8 9.70.2 1.0 0.8 1.5 Chloramphenicol Ribosomal Antibacterialpeptidyltransferase inhibitor 0.9 0.8 0.7 0.6 0.8 0.8 1.1 Naloxonehydrochloride Opiate antagonist Opioate antidote 1.0 0.8 2.2 0.7 0.9 0.81.2 Epirizole Antiinflammatory 0.8 0.8 0.7 0.6 0.8 0.7 1.0 Metolazone —Diuretic 1.0 0.9 0.7 0.6 0.8 0.7 1.1 Diprophylline Phosphodiesteraseinhibitor Cardiac analeptic 33.6 1.2 1.5 0.8 2.2 29.7 4.7 Ciprofloxacinhydrochloride Anti-bacterial 0.6 0.7 0.6 0.5 1.1 0.6 1.2 Triamterene Noncompetitive Diuretic aldosterone antagonist 12.2 1.1 0.5 0.6 1.6 0.7 3.5Ampicillin trihydrate Antibacterial 0.8 0.9 0.6 0.7 1.3 0.8 1.1 DapsoneFolic acid antagonist Antiinflammatory 0.9 0.9 0.8 0.6 1.0 0.9 1.1Haloperidol Dopamine antagonist Antipsychotic 0.9 1.0 6.5 0.6 0.8 0.71.1 Troleandomycin Ribosomal protein synthesis Antibacterial inhibitor0.9 1.0 0.7 0.6 0.9 0.8 1.3 Naltrexone hydrochloride Opioid antagonistAnalgesic dihydrate 1.4 1.0 0.7 0.7 2.5 0.9 1.3 PyrimethamineAntimalarial 1.0 0.9 0.8 0.7 0.9 0.8 1.1 Chlorpheniramine maleate H1antagonist Antihistaminic 0.9 0.9 0.7 0.7 0.9 0.8 1.2 Hexamethoniumdibromide Ganglion blocking agent Antihypertensor dihydrate 0.9 0.9 0.70.7 0.9 0.8 1.2 Nalbuphine hydrochloride Opioid ligand Analgesic 0.7 0.90.6 0.6 0.7 0.6 1.2 Diflunisal Cyclooxygenase inhibitor Antiinflammatory0.9 0.9 0.7 0.8 0.9 0.8 1.2 Picotamide monohydrate Eicosenoid receptorAntithrombotic antagonist 0.1 0.1 0.4 0.1 0.2 0.1 0.5 NiclosamideHelmintic DNA damage Anthelmintic 1.0 1.0 0.8 0.7 0.9 0.7 1.2Triamcinolone — Antiinflamatory 1.0 1.0 0.9 0.7 0.9 0.9 1.0 Midodrinehydrochloride Alpha adrenergic Hypotensor 0.9 0.9 0.7 0.7 0.9 1.0 1.2Vincamine — Cerebral antianoxic 1.0 1.0 1.0 0.8 0.9 0.9 1.1 Thalidomide— Immunosuppressant 0.9 0.9 0.6 0.8 1.0 0.9 1.2 IndomethacinCyclooxygenase inhibitor Antiinflammatory 13.1 1.1 1.0 1.6 2.3 1.0 4.3Oxolinic acid Topoisomerase II inhibitor Antibacterial 0.9 0.9 0.8 0.81.0 1.0 1.2 Cortisone Antiinflammatory 0.9 0.8 0.7 0.7 0.9 0.9 1.4Nimesulide Cyclooxygenase 2 inhibitor Antiinflammatory 1.0 1.0 0.8 0.71.0 1.0 1.3 Prednisolone — Glucocorticoid 1.0 0.9 0.7 0.8 0.9 0.9 1.2Hydrastinine hydrochloride Dopamine receptor blocker Cardiotonic 0.8 0.80.8 0.6 0.7 0.8 1.2 Fenofibrate Lipoprotein lipase activatorHypolipidemiant 0.8 0.9 0.8 0.7 0.9 0.8 1.1 PentoxifyllinePhosphodiesterase inhibitor Vasodilatator 0.9 0.9 0.7 0.7 0.8 0.8 1.3Bumetanide Vascular cyclooxygenase Diuretic activator 1.0 1.0 0.7 0.80.9 0.8 1.3 Metaraminol bitartrate Adrenergic agonist Vasoconstrictor0.9 0.9 0.8 0.7 0.9 0.9 1.3 Labetalol hydrochloride 0.9 1.0 0.8 0.7 0.90.8 1.4 Salbutamol Beta adrenergic agonist Bronchodilatator 0.9 1.0 1.00.8 2.3 0.9 1.2 Cinnarizine H1 antagonist Antihistaminic 0.9 0.9 0.8 0.70.9 0.9 1.3 Prilocaine hydrochloride Na+ channel blocker Localanesthesic 0.8 1.0 0.8 0.7 0.9 0.9 1.3 Methylprednisolone, 6-alphaGlucocorticoid 0.9 0.9 0.7 0.8 0.8 0.8 1.1 Camptothecine (S,+)Topoisomerase I inhibitor Antitumor agent 1.0 1.0 0.9 0.8 0.9 0.9 1.1Quinidine hydrochloride Heme polymerase inhibitor Antiarrhythmicmonohydrate 1.2 0.9 1.2 1.0 1.1 1.0 1.4 Procaine hydrochloride Na+channel blocker Local anesthesic 1.3 0.9 0.6 1.1 1.4 1.1 1.4Bromocryptine mesylate Prolactin inhibitor Antiparkinsonian 1.2 0.9 1.11.0 1.1 1.1 1.6 Moxisylyte hydrochoride Alpha antagonist Vasodilatator1.3 1.0 1.2 1.0 1.1 1.1 1.3 Metanephrine hydrochloride DL Isoprenalineuptake inhibitor extraneuronal 1.3 1.0 1.2 0.9 1.2 1.0 1.4 Betazolehydrochloride Histamine analog Gastric secretion stimulant 1.3 1.0 1.00.8 1.0 1.0 1.4 Dehydrocholic acid — Choleretic 1.2 1.0 1.0 1.0 1.0 1.01.4 Isoxicam Cyclooxygenase inhibitor Antiinflammatory 1.1 0.9 0.5 0.91.1 0.9 1.3 Hesperetin P450 inhibitor 1.2 1.0 0.8 0.9 1.1 1.0 1.4Naproxen Cyclooxygenase inhibitor Antiinflammatory 1.5 1.2 0.9 1.3 1.61.1 1.4 Perphenazine Dopamine antagonist Antipsychotic 1.3 1.1 1.0 0.81.1 1.0 1.5 Naphazoline hydrochloride Adrenergic ligand Vasoconstrictor1.3 1.0 3.2 1.3 2.6 1.1 1.2 Mefloquine hydrochloride Heme polymeraseinhibitor Antimalarial 1.3 1.0 0.8 1.0 1.1 1.1 2.0 Ticlopidinehydrochloride ADP antagonist Platelet antiaggregant 1.8 1.8 5.2 1.4 5.61.3 1.7 Isoconazole Sterol 14-demethylase Antibacterial inhibitor 1.21.1 1.0 0.9 1.3 1.2 1.5 Dicyclomine hydrochloride AnticholinergicAntispasmodic 1.4 1.0 1.2 1.0 1.1 1.0 1.3 Spironolactone Aldosteroneantagonist Diuretic 1.4 1.1 1.1 0.8 1.1 0.9 1.2 Amyleine hydrochlorideNa+ channel blocker Local Anesthesic 1.4 1.1 1.0 1.0 1.1 1.0 1.4Pirenzepine dihydrochloride M1 antagonist Antiulcerative 1.3 1.2 1.1 1.11.1 1.1 1.3 Lidocaine hydrochloride Na+ channel blocker Local anesthesic1.4 1.2 1.2 1.1 1.1 1.0 1.2 Dexamethasone acetate Antiinflammatory 1.41.1 1.2 0.9 1.1 1.3 1.5 Ranitidine hydrochloride H2 antagonistAntiulcerative 1.3 1.1 1.0 1.0 1.2 1.2 1.4 Fludrocortisone acetate —Mineralocorticoid 1.1 1.3 0.4 1.0 1.1 1.2 1.5 Tiratricol, 3,3′,5- —Hypocholesterolemic drug triiodothyroacetic acid 1.3 1.1 1.0 0.9 1.3 1.11.4 Fenoterol hydrobromide Beta adrenergic agonist Bronchodilatator 1.01.0 0.5 1.0 1.3 1.1 1.8 Flufenamic acid Cyclooxygenase inhibitorAnalgesic 1.5 1.2 1.0 1.1 1.2 1.2 1.8 Homochlorcyclizine H1 antagonistAntihistaminic dihydrochloride 13.6 1.5 2.2 2.2 2.9 1.4 24.4 FlumequineTopoisomerase II inhibitor Antibacterial 1.3 1.2 1.1 0.9 1.1 1.1 1.7Diethylcarbamazine citrate Lipoxygenase inhibitor Antihelmintic 1.0 1.10.5 0.8 1.4 1.2 1.6 Tolfenamic acid Cyclooxygenase inhibitorAntiinflammatory 1.2 1.0 1.7 0.8 1.0 1.2 1.5 Chenodiol DetergentAnticholelithogenic 1.1 1.2 0.6 1.0 1.2 1.2 1.4 Meclofenamic acid sodiumsalt Cyclooxygenase inhibitor Antiinflammatory monohydrate 1.8 1.2 0.81.6 3.4 1.4 1.5 Perhexiline maleate Ca2+ blocking agent Vasodilatator1.1 1.1 1.0 1.0 1.2 1.0 1.4 Kawain Ca++ channel blocker Antiaggregant1.4 1.1 1.2 0.9 1.1 1.0 1.6 Oxybutynin chloride AnticholinergicSpasmolytic 1.2 1.1 1.0 0.8 3.1 1.1 7.2 Trimethoprim Folic acidantagonist Antibacterial 1.3 1.2 1.0 0.9 1.2 1.2 1.3 Spiperone D2antagonist Antipsychotic 1.3 1.2 0.9 0.9 1.2 1.1 1.6 Metoclopramide5-HT3 antagonist Antiemetic monohydrochloride 1.4 1.2 1.4 1.0 1.1 1.11.4 Pyrilamine maleate H1 antagonist Antihistaminic 0.9 0.9 0.6 0.7 0.90.9 1.3 Fenbendazole Microtubule formation Antihelmintic inhibitor 1.31.1 3.5 1.0 1.2 1.0 1.2 Sulfinpyrazone — Uricosuric 1.2 0.8 1.1 1.1 0.91.0 1.5 Trichlorfon Cholinesterase inhibitor Antihelminthic 1.2 0.9 1.20.8 1.0 1.0 1.3 Glipizide 1.0 0.8 1.0 0.9 0.9 1.0 1.5 CarbamazepineCholinergic antagonist Anticonvulsivant 1.0 0.9 1.1 1.0 1.0 0.9 1.3Loxapine succinate Dopamine antagonist Anxiolytic 1.1 0.9 1.1 1.0 1.51.0 1.6 Triflupromazine hydrochloride Dopaminergic antagonist ?Antipsychotic 1.0 0.9 0.9 0.8 1.0 1.0 1.6 Hydroxyzine dihydrochloride H1antogonist Antihistaminic 0.9 0.9 0.7 0.8 1.2 0.9 1.6 Mefenamic acidCyclooxygenase inhibitor Antiinflammatory 1.0 0.9 0.8 1.0 1.0 0.9 1.5Diltiazem hydrochloride Ca2+ channel inhibitor (L- Antianginal Type) 1.20.9 1.1 0.8 1.1 1.2 1.7 Acetohexamide Blocking of ATP-sensitive K+Antidiabetic (type II, channel noninsulin-dependent) 1.1 0.8 0.9 0.8 1.01.0 1.3 Methotrexate 1.0 1.0 1.1 1.0 1.0 1.1 1.6 Sulpiride D2 antagonistAntipsychotic 1.3 1.0 2.2 1.1 1.6 1.0 1.6 Astemizole H1 antagonistAntihistaminic 1.3 0.9 1.0 1.0 1.1 0.9 1.4 Benoxinate hydrochloride Na+channel blocker Local anesthesic 0.8 1.2 8.4 0.5 1.0 1.1 1.7 Clindamycinhydrochloride Ribosomal protein synthesis Antibacterial inhibitor 1.41.0 1.0 1.5 4.2 1.0 1.5 Oxethazaine Na+ channel blocker Local anesthesic1.7 1.5 ### 1.1 3.1 1.0 1.2 Terfenadine H1 antagonist Antihistaminic 1.41.0 1.0 1.0 1.0 1.0 1.7 Pheniramine maleate H1 antagonist Antihistaminic21.9 1.0 2.9 5.5 2.2 10.0 22.0 Cefotaxime sodium salt Bacterialtranspeptidase Antibacterial inhibitor 1.4 1.0 1.2 1.0 1.3 1.1 1.5Tolazoline hydrochloride Alpha antagonist Vasodilatator 0.8 1.0 9.2 0.24.4 0.5 1.0 Tetracycline hydrochloride Ribosomal protein synthesisAntibacterial inhibitor 1.2 1.1 1.4 1.0 1.0 1.2 1.4 PiroxicamCyclooxygenase inhibitor Antiinflammatory 1.0 1.0 1.1 0.9 1.1 1.2 1.5Dantrolene sodium salt Blocker of Ca2+ release Skeletal muscle relaxant1.3 1.1 1.2 0.9 1.1 1.2 1.4 Pyrantel tartrate Neuromuscular depolarizingAnthelmintic agent 0.9 1.0 0.9 1.0 1.0 1.2 1.3 Trazodone hydrochloride5-HT uptake inhibitor Antidepressant 1.3 1.0 1.1 1.0 1.2 1.1 1.6Fenspiride hydrochloride Bradykinin antagonist Antiinflammatory 1.2 1.11.1 1.0 1.1 1.2 1.5 Glafenine hydrochloride — Analgesic 1.3 1.0 0.9 0.91.3 1.2 1.5 Gemfibrozil — Antihyperlipoproteinemic 1.4 1.1 0.9 1.2 1.11.1 1.4 Pimethixene maleate Anticholinergic Antihistaminic 1.2 1.0 0.90.9 1.0 1.1 1.3 Mefexamide hydrochloride — Psychoanaleptic 1.3 1.0 1.00.8 1.3 1.0 1.4 Pergolide mesylate Dopaminergic agonist Antiparkinsonian1.4 1.1 1.0 0.9 1.1 1.2 1.5 Tiapride hydrochloride Dopamine antagonistAntidyskinetic 1.2 1.0 1.1 0.9 1.1 1.1 1.3 Acemetacin Cyclooxygenaseinhibitor Antiinflammatory 0.8 0.9 0.9 0.8 0.7 0.8 1.2 Mebendazole 1.31.1 1.1 1.0 1.0 1.2 1.2 Benzydamine hydrochloride 5-HT receptorantagonist Analgesic 0.8 0.8 0.7 0.8 1.0 0.8 1.6 Fenbufen Cyclooxygenaseinhibitor Antiinflammatory 1.3 1.1 0.9 1.1 1.0 1.0 1.3 Fipexidehydrochloride Glutamatergic Nootropic 1.1 0.9 1.0 1.1 1.0 1.0 1.5Ketoprofen Cyclooxygenase inhibitor Antiinflammatory 1.3 1.1 0.7 0.9 1.11.0 1.4 Mifepristone Progesterone receptor Abortifacient antagonist 1.11.1 1.0 1.1 1.0 1.1 1.7 Indapamide Diuretic 1.4 1.1 1.2 0.9 1.1 1.1 1.4Diperodon hydrochloride — Local anesthesic 1.4 0.8 1.2 0.9 1.1 1.2 1.3Morantel tartrate Fumarate reductase Anthelmintic inhibitor 1.3 0.9 1.20.9 1.5 1.2 1.2 Verapamyl hydrochloride Alpha1 antagonistAntyhypertensive 1.4 0.8 1.1 0.9 1.3 1.1 1.4 Homatropine hydrobromide(R,S) Muscarinic antagonist Antispasmodic 1.3 0.9 0.8 0.9 1.2 1.1 1.4Dipyridamole 1.4 0.9 0.5 0.9 1.5 1.0 1.4 Nifedipine L-type Ca2+ channelsblocker Antihypertensor 50.1 4.0 ### 18.8 12.3 23.8 1.8 ChlorhexidineDetergent Bacteriostatic 1.4 1.0 1.4 1.1 1.5 1.1 1.4 Chlorpromazinehydrochloride Dopamine antagonist Antiemetic 1.4 0.9 1.0 0.9 1.5 1.2 1.5Loperamide hydrochloride Ca2+ channel antagonist Antidiarrhoeic 1.3 0.91.1 1.0 1.3 1.0 1.4 Diphenhydramine hydrochloride H1 receptor antagonistAntihistaminic 0.8 0.9 6.8 0.2 3.9 0.7 1.0 Chlortetracyclinehydrochloride Ribosomal protein synthesis Antibacterial inhibitor 1.30.8 1.0 0.9 1.3 1.1 1.3 Minaprine dihydrochloride Dopamine agonistAntidepressant 1.9 4.0 2.1 2.1 6.6 1.4 1.3 Tamoxifen citrate Oestrogenreceptor antagonist 1.4 2.5 2.2 1.4 4.7 1.2 1.6 Miconazole Sterol14-demethylase Antifungal inhibitor 1.3 1.0 1.4 1.0 1.4 1.2 1.2Nicergoline Alpha agonist Vasodilatator 1.1 1.0 1.1 1.0 1.5 1.1 1.5Isoxsuprine hydrochloride beta adrenergic agonist Vasodilatator 1.4 1.01.1 0.9 1.5 1.3 1.2 Canrenoic acid potassium salt DetergentAntihypercholesterolemic 1.3 1.1 1.1 0.9 1.3 1.1 1.2 Acebutololhydrochloride Beta1 antagonist Antianginal 1.5 1.0 1.0 1.1 1.7 1.1 1.3Thioproperazine dimesylate Dopamine antagonist Antipsychotic 1.1 1.0 1.30.9 1.5 1.0 1.2 Tolnaftate — Antifungal 1.3 1.0 1.0 0.9 1.5 1.2 1.3Dihydroergotamine tartrate Serotonine antagonist Antimigraine 33.1 1.51.8 0.8 3.1 31.3 3.1 Norfloxacin Topoisomerase II inhibitorAntibacterial 1.2 1.0 4.3 1.0 1.3 1.2 1.5 Lisinopril Converting enzymeinhibitor Antihypertensor 1.3 1.1 1.0 1.0 1.4 1.1 1.3 Antimycin AInhibitor of mitochondrial Antifungal electron transport 0.9 1.3 7.9 0.91.2 1.2 1.3 Lincomycin hydrochloride Ribosomal protein synthesisAntibacterial inhibitor 1.2 1.0 1.2 0.9 1.3 1.2 1.4 Xylometazolinehydrochloride Vasoconstrictor 1.5 1.1 1.3 1.0 1.6 1.2 1.4 Telenzepinedihydrochloride M1 muscarinic antagonist Antiulcerative 1.1 1.0 1.1 1.11.4 1.2 1.3 Oxymetazoline hydrochloride Partial alpha2A agonistVasoconstrictor 1.3 1.5 3.7 1.3 6.7 1.3 1.3 Econazole nitrate Ergosterolsynthesis Antifungal inhibition 1.2 1.0 1.3 1.0 1.6 1.2 1.2 NifenazoneCyclooxygenase inhibitor Analgesic 1.3 1.0 1.1 1.0 1.4 1.2 1.5Bupivacaine hydrochloride Na+ channel blocker Local anesthesic 1.3 1.11.4 0.9 1.4 1.3 1.3 Griseofulvin Enzymatic inductor Antifungal 1.4 1.01.2 1.1 1.5 1.2 1.2 Clemastine fumarate H1 antagonist Antihistaminic 1.21.2 1.6 1.0 1.6 1.2 1.4 Clemizole hydrochloride H1 antagonistAntihistaminic 1.0 0.9 3.4 0.2 5.3 0.5 1.1 Oxytetracycline dihydrateRibosomal protein synthesis Antibacterial inhibitor 1.2 1.0 1.1 1.1 1.51.2 1.4 Tropicamide Muscarinic antagonist Mydriatic 1.7 1.3 2.3 1.3 8.71.3 1.2 Pimozide Dopamine antagonist ? Antipsychotic 1.2 1.0 1.1 0.9 1.41.2 1.4 Nefopam hydrochloride — Analgesic 1.2 1.1 1.3 0.9 1.5 1.2 1.4Amodiaquin dihydrochloride Heme polymerase inhibitor Antimalarialdihydrate 1.4 1.0 1.1 1.1 1.4 1.2 1.2 Phentolamine hydrochloride Alphaadrenergic antagonist Antihypertensor 1.3 1.0 1.1 1.0 1.4 1.1 1.3Mebeverine hydrochloride Antispasmodic 1.3 0.8 1.3 1.0 1.1 1.1 1.7Todralazine hydrochloride — Antihypertensor 1.0 1.0 8.2 1.0 1.5 0.7 1.6Erythromycin Ribosomal protein synthesis Antibacterial inhibitor 1.2 0.91.2 1.0 1.3 1.1 1.7 Imipramine hydrochloride 5-HT transport inhibitorAntidepressant 1.1 1.0 3.6 1.0 1.1 1.0 1.6 Oleandomycin phosphateRibosomal protein synthesis Antibacterial inhibitor 1.1 0.9 0.8 1.0 1.10.8 1.5 Sulindac Cyclooxygenase inhibitor Antiinflammatory 4.7 0.9 1.20.9 1.5 1.0 4.2 Didanosine Transrcriptase inverse Antiviral inhibitor1.2 1.0 1.4 1.1 1.3 1.0 1.8 Amitryptiline hydrochloride Alpha 1antogonist Antidiabetic 1.3 1.0 8.4 1.0 1.1 0.9 1.4 Josamycin Ribosomalprotein synthesis Antibacterial inhibitor 1.2 1.0 1.3 1.0 1.3 0.9 1.5Adiphenine hydrochloride Anticholinergic Local anesthesic 1.1 1.0 1.01.0 1.3 1.0 1.6 Paclitaxel Tubuline inhibitor Antineoplastic 1.3 0.9 1.31.1 1.3 1.1 1.7 Dibucaine Na+ channel blocker Local anesthesic 1.3 1.22.1 1.0 1.5 1.0 1.6 Ivermectin GABA ligand Anthelmintic 1.2 0.9 1.1 0.91.2 0.9 1.4 Prednisone — Glucocorticoid 1.3 1.0 1.1 1.1 1.2 1.0 1.5Gallamine triethiodide M2 antagonist allosteric Muscle relaxant 1.6 1.01.3 1.3 2.9 1.1 1.5 Thioridazine hydrochloride Ca2+ channel antagonistNeuroleptic 1.2 1.0 1.1 1.0 1.4 0.9 1.3 Neomycin sulfate Ribosomalprotein synthesis Antibacterial inhibitor 1.2 1.1 1.5 1.1 1.3 1.0 1.6Diphemanil methylsulfate Anticholinergic Bronchodilatator 1.2 1.0 1.20.7 1.2 0.9 1.3 Dihydrostreptomycin sulfate Ribosomal protein synthesisAntibacterial inhibitor 1.2 1.1 1.2 1.1 1.2 1.0 1.3 Trimethobenzamide D2antagonist Antiemetic hydrochloride 1.3 1.0 1.1 1.0 1.5 0.9 1.4Gentamicine sulfate Ribosomal protein synthesis Antibacterial inhibitor1.2 1.1 1.0 1.3 1.4 1.5 1.2 Etodolac Cyclooxygenase inhibitorAntiinflammatory 1.2 1.0 1.1 1.3 1.4 1.2 1.3 Ifenprodil tartrateAdrenergic antagonist Vasodilatator 1.2 1.1 1.4 1.2 1.4 1.3 1.5Scopolamin-N-oxide Anticholinergic Antiparkinsonian hydrobromide 1.7 1.31.1 1.4 2.6 1.3 1.6 Flunarizine dihydrochloride Na+ channel blockerVasodilatator 1.3 1.0 1.2 1.2 1.4 1.2 1.5 Hyoscyamine (L) CholinergicAntispasmodic 1.7 1.1 1.2 1.5 3.4 1.4 1.3 Trifluoperazinedihydrochloride Dopamine antagonist Psycholeptic 1.2 0.9 1.0 1.1 1.3 1.11.4 Chlorphensin carbamate — Muscle relaxant 1.2 1.1 1.3 1.2 1.3 1.2 1.3Enalapril maleate Converting enzyme inhibitor Antihypertensor 16.8 1.61.3 1.0 1.1 1.3 4.3 Metampicillin sodium salt Bacterial transpeptidaseAntibacterial inhibitor 2.3 1.1 6.9 0.6 3.9 0.7 0.9 Minocyclinehydrochloride Ribosomal protein synthesis Antibacterial inhibitor 1.51.1 1.2 1.4 1.4 1.3 1.1 Dilazep dihydrochloride Adenosine uptakeinhibitor Vasodilatator 1.2 1.0 0.9 1.1 1.3 1.2 1.4 GlibenclamideATP-dependent K+ channel Antidiabetic inhibitor 15.7 1.4 2.8 1.0 3.720.0 4.1 Ofloxacin Topoisomerase II inhibitor Antibacterial 1.4 1.1 1.11.1 1.3 1.4 1.4 Guanethidine sulfate Catecholamine depletorAntihypertensor 26.8 1.5 3.4 1.0 3.8 30.2 3.5 Lomefloxacin hydrochlorideTopoisomerase II inhibitor Antibacterial 1.2 1.0 1.2 1.0 1.3 0.9 1.4Quinacrine dihydrochloride Monoamine oxydase Antimalarial dihydrateinhibitor 1.3 1.0 1.0 1.2 1.3 1.1 1.2 Orphenadrine hydrochloride H1antogonist Antihistaminic 1.5 1.4 1.2 1.5 1.6 1.3 1.2 Clofilium tosylateK+ channel blocker Antiarrhythmic 1.3 1.0 1.1 1.2 1.3 1.1 1.5 ProglumideGastrin inhibitor Antiulcerative 1.3 1.1 1.0 1.5 1.6 1.1 1.6Fluphenazine dihydrochloride Dopamine antagonist Antipsychotic 0.5 0.90.4 0.3 0.4 0.4 0.3 Streptomycin sulfate Ribosomal protein synthesisAntibacterial inhibitor 0.5 0.9 0.3 0.4 0.5 0.4 0.3 Testosteronepropionate — Androgen 0.5 0.9 0.3 0.4 0.4 0.4 0.4 Alfuzosinhydrochloride Alpha 1-adrenergic Antihypertensor antagonist 0.4 0.9 0.40.4 0.4 0.4 0.3 Arecoline hydrobromide Cholinergic Anthelmintic 0.4 0.90.4 0.4 0.3 0.3 0.3 Chlorpropamide Glucagon secretogen, Antidiabeticsomatostatin secretogen 0.5 1.0 0.2 0.4 0.4 0.4 0.3 Thyroxine (L)Thyroid hormone Hypocholesterolemic drug 0.5 0.8 0.3 0.5 0.4 0.4 0.4Phenylpropanolamine Alpha adrenergic agonist Decongestant hydrochloride0.6 0.9 0.4 0.4 0.4 0.4 0.4 Tocopherol (R,S) — Anti-oxidant 0.5 0.9 0.40.5 0.4 0.3 0.4 Ascorbic acid — Vitamin 0.5 0.8 0.4 0.5 0.4 0.4 0.4Pepstatin A Aspartic proteases Antiviral irreversible inhibitor 0.6 0.90.4 0.6 0.5 0.4 0.5 Methyldopa (L,−) L-aromatic aminoacidAntihypertensor decarboxylase inhibitor 0.5 0.8 0.4 0.5 0.5 0.4 0.4SR-95639A M1 agonist receptor 16.5 0.9 0.7 2.1 0.7 5.2 12.0 Cefoperazonedihydrate Bacterial transpeptidase Antibacterial inhibitor 0.5 0.9 0.40.5 0.5 0.4 0.4 Adamantamine fumarate Inhibition of viral uncoatingAntiviral and viral assembly, alteration of dopamine release andreuptake 0.7 0.8 0.4 0.6 0.4 0.4 0.4 Zoxazolamine Uric acid uptakeinhibitor Muscle relaxant 0.7 3.3 0.6 0.7 2.0 0.5 0.5 Butoconazolenitrate Ergosterol inhibitor Antifungal 0.5 0.9 0.5 0.4 0.5 0.5 0.4Tacrine hydrochloride hydrate Cholinesterase inhibitor Cognitionenhancer 0.7 1.7 0.7 0.6 2.2 0.5 0.5 Amiodarone hydrochloride Na+channel blocker, K+ Antiarrhythmic channel blocker, non- competitivebeta-adrenergic blocker 0.5 0.9 0.6 0.5 0.5 0.5 0.6 Bisoprolol fumarateBeta1 antagonist Antihypertensor 0.5 0.9 0.5 0.7 0.5 0.4 0.5Amphotericin B Ergosterol ligand Antibacterial 0.8 0.9 0.6 0.6 0.6 0.60.6 Serotonin hydrochloride 5-HT agonist Neurotransmitter 0.7 0.9 0.50.6 0.7 0.6 0.6 Tubocurarine chloride Curarising Muscle relaxantpentahydrate (+) 23.9 0.9 2.3 7.8 0.9 0.6 15.6 Cefotiam hydrochlorideAntibacterial 0.7 0.9 0.5 0.6 0.7 0.6 0.6 Dihydroergocristine mesylate5-HT antagonist, partial Vasodilatator adrenergic agonist, partialdopaminergic agonist 0.6 0.9 0.4 0.5 0.5 0.6 0.6 Azathymine, 6Antimetabolite Anticancer 0.6 0.9 0.5 0.5 0.6 0.5 0.6 NoscapineInhibitor of carbachol- Antitussive stimulated phosphoinositide turnover0.6 1.0 0.5 0.5 0.5 0.6 0.6 Benperidol Dopamine antagonist, 5-HTAntipsychotic antagonist 0.6 0.9 0.3 0.5 0.6 0.5 0.6 SyrosingopineCatecholamine depletor — 10.7 0.9 0.5 2.5 1.5 0.5 0.7 Cefaclor Bacterialtranspeptidase Antibacterial inhibitor 0.6 0.8 0.5 0.5 0.5 0.5 0.5Atropine sulfate monohydrate Muscarinic antagonist Anticholinergic 42.21.0 0.6 24.1 10.0 23.6 0.7 Colistin sulfate Performs membraneAntibacterial ionophores 0.7 0.9 0.4 0.5 0.5 0.6 0.7 Eserine sulfate,physostigmine Cholinesterase inhibitor Ophtalmic agent sulfate 0.9 2.32.1 0.5 0.6 0.5 0.6 Daunorubicin hydrochloride DNA intercalingAntineoplastic 0.6 0.9 0.5 0.5 0.6 0.5 0.6 Aconitine Open TTX-Na+channel Analgesic 0.6 0.9 0.5 0.5 0.5 0.5 0.5 Dosulepin hydrochloride —Antidepressant 0.7 1.0 0.3 0.6 1.1 0.4 0.6 Rescinnamin Catecholaminedepletor Antihypertensor 11.5 0.9 0.4 3.2 0.5 7.8 12.9 Ceftazidimepentahydrate Antibacterial 0.6 0.8 0.4 0.5 0.5 0.4 0.6 Dihydroergotoxinemesylate High affinity GABA A Anticonvulsant receptor Cl− channel,prolactin inhibitor 0.5 0.9 0.5 0.5 0.5 0.4 0.6 Iobenguane sulfate —Antineoplastic if radiolabeled 0.5 0.9 0.5 0.6 0.5 0.5 0.5 Emetinedihydrochloride Protein synthesis inhibitor, Antiamebic 5-HT ligand 0.71.0 0.5 0.6 0.7 0.6 0.7 Tremorine dihydrochloride Cholinergic Convulsant0.6 1.0 0.5 0.5 0.7 0.6 0.7 Androsterone — Antihypertensor 0.7 0.9 0.60.5 0.7 0.6 0.8 Practolol Beta antagonist Antihypertensor 0.7 1.0 0.70.5 0.7 0.7 0.7 Anisomycin Acetylcholine esterase Antiprotozoalinhibitor 2.1 0.9 0.6 1.2 1.8 0.6 4.2 Zidovudine, AZT Transcriptasereverse Antiviral inhibitor 0.6 1.0 0.6 0.5 0.6 0.6 0.7 CarbarsoneAntiamebic 0.7 0.9 0.5 0.5 1.0 0.6 0.7 Sulfisoxazole Inhibitor of folicacid Antibacterial biosynthesis, Eta endothelin receptor antagonist 0.20.3 0.0 0.2 0.2 0.2 0.5 Apigenin MAP kinase inhibitor Antiproliferative0.7 1.0 0.6 0.5 0.6 0.7 0.8 Zaprinast cGMP phosphodiesterase Erectogeninhibitor, phosphodiesterase 5 inhibitor 0.6 0.9 0.5 0.5 0.6 0.6 0.7Aspartic acid, N-acetyl (R,S) — — 0.7 1.0 0.7 0.6 0.7 0.6 0.7Chlormezanone — Skeletal muscle relaxant 11.9 1.1 0.5 0.6 1.3 0.6 0.6Bacampicillin hydrochloride Bacterial transpeptidase Antibacterialinhibitor 0.6 1.0 0.6 0.6 0.7 0.6 0.9 Procainamide hydrochloride Alphaantagonist, Antiarrhythmic antinuclear antibodies, anticholinergic 0.60.9 0.1 0.5 0.6 0.6 0.7 Betulinic acid Apoptosis inducer, PLA2Antimalarial inhibitor 0.6 0.9 0.5 0.5 0.6 0.6 0.7 N6-methyladenosineAntimetabolite Anticancer 0.6 0.9 1.4 0.6 0.8 0.7 0.8 Biotin — Vitamin H0.8 1.1 0.6 0.5 0.7 0.7 0.9 Guanfacine hydrochloride Alpha 2A agonistAntihypertensor 0.6 1.0 0.5 0.6 0.8 0.6 0.9 Bisacodyl Na+ uptakeinhibitor Cathartic 0.6 0.9 0.8 0.6 0.7 0.6 0.8 Domperidone DopamineAntagonists Antiemetic 0.8 0.9 0.5 0.7 0.7 0.7 0.7 Calciferol Stimulatorof calcium and Vitamin D phosphate absorption 0.9 1.0 0.7 0.8 1.0 0.80.8 Metixene hydrochloride Anticholinergic Antiparkinsonian 0.9 0.9 0.60.6 0.8 0.8 0.9 Tetracaine hydrochloride Local anesthesic 0.7 1.0 0.50.2 1.0 0.8 0.8 Nitrofural Bacterial DNA damage Antibacterial 0.9 1.00.4 0.7 0.9 0.6 0.9 Mometasone furoate — Antiinflammatory 0.7 0.8 0.20.6 0.8 0.6 0.8 Omeprazole Non competitive ATPase H+ Antiulcerative pumpinhibitor 0.7 1.2 0.5 0.6 0.7 0.7 1.0 Tomatidine Cholinesterase activityAntifungal 0.7 1.0 0.7 0.6 0.8 0.7 1.1 Propylthiouracil AntimetaboliteAntihyperthyroid 2.0 1.1 0.7 3.8 1.6 0.9 0.8 Dacarbazine Alkylatingagent Antineoplastic 0.9 1.0 0.6 0.7 0.8 0.8 0.9 Terconazole Sterol14-demethylase Antifungal inhibitor 0.6 0.9 0.7 0.6 0.6 0.8 0.8Ipratropium bromide Antimuscarinic agent Bronchodilatator 0.6 0.8 0.50.5 0.7 0.6 0.8 Tiaprofenic acid Cyclooxygenase inhibitorAntiinflammatory 0.8 1.0 0.8 0.6 0.8 0.7 0.8 Acetopromazine maleate saltDopaminergic antagonist ? Tranquilizer 1.2 1.5 ### 0.8 1.1 0.9 0.9Vancomycin hydrochloride Bacterial mucopeptide Antibacterialbiosynthesis inhibitor 0.7 1.0 0.7 0.6 0.8 0.6 0.8 Rauwolscinehydrochloride Alpha2 antagonist Antidepressant 0.5 0.9 0.5 0.6 0.7 0.60.7 Artemisinin Oxidant Antimalarial 0.7 1.0 0.7 0.6 0.6 0.7 0.8Corynanthine hydrochloride Alpha1-adrenoreceptor Anti-leishmania drugantagonist 1.0 1.0 0.7 0.6 0.7 0.7 0.8 Propafenone hydrochloride Betaadrenergic antagonist Antiarrhythmic 0.7 1.0 0.8 0.5 0.7 0.7 0.9Palmatine chloride Anticholinestrase activity Uterine contractant 0.61.0 0.6 0.6 0.9 0.6 0.9 Ethamivan — Respiratory analeptic 0.8 1.2 0.60.7 1.0 0.7 0.8 Trimethylcolchicinic acid Tubuline inhibitor ?Anticancer agent 1.0 0.9 0.7 0.7 0.8 0.9 0.8 Furosemide Na+ Cl− uptakeinhibitor, Diuretic carbonic anhydrase inhibitor 1.9 5.4 2.0 2.5 4.1 6.80.9 Suloctidil — Vasodilatator 0.9 1.0 0.5 0.6 0.9 0.9 0.8 Methapyrilenehydrochloride Histamine H1 antagonist Antihistaminic 0.8 0.9 0.7 0.6 0.80.8 0.8 Carcinine — Anti-oxidant 0.9 0.8 0.7 0.7 0.7 0.9 0.8 Desipraminehydrochloride Adrenergic transport Antidepressant inhibitor, 5-HTtransport inhibitor 0.9 0.9 0.7 0.6 0.8 0.9 0.8 Carisoprodol — Musclerelaxant 0.7 0.7 0.5 0.5 0.6 0.7 0.8 Clorgyline hydrochloride Monoamineoxidase A Antidepressant inhibitor 0.9 0.9 0.6 0.7 0.8 0.8 0.8Cephalosporanic acid, 7-amino Bacterial transpeptidase Antibacterialinhibitor 0.8 0.9 0.7 0.7 0.8 0.8 0.9 Clenbuterol hydrochloride Betaadrenergic agonist Bronchodilatator 0.5 0.5 0.0 0.3 0.4 0.4 0.9 Chicagosky blue 6B Competitive glutamate — uptake inhibitor 0.8 0.8 1.1 0.6 0.80.9 0.9 Maprotiline hydrochloride Noradrenaline uptake Antidepressantinhibitor, 5-HT uptake inhibitor 0.8 0.8 0.6 0.6 0.7 0.8 0.9 Buflomedilhydrochloride — Vasodilatator 0.8 0.9 0.5 0.7 0.9 0.8 0.9 ThioguanosineAntimetabolite Anticancer 0.8 0.9 0.5 0.7 0.8 0.7 0.8 Chlorogenic acidGlucose-6-phosphate Antiinflammatory translocase inhibitor 1.0 1.0 0.60.8 1.2 0.9 1.0 Chlorprothixene hydrochloride D2 dopamine receptorNeuroleptic antagonist, GABAA receptors antagonist 0.8 1.0 0.6 0.7 0.80.7 0.9 Roxatidine Acetate HCl Antiulcerative 0.8 0.9 0.5 0.7 0.8 0.90.9 Ritodrine hydrochloride Beta2 agonist Tocolytic 1.0 0.9 0.7 0.7 0.80.9 1.0 Cholecalciferol — Vitamin 0.9 0.9 0.7 0.8 0.9 1.0 1.0 Clozapine5-HT antagonist, dopamine Antipsychotic antagonist, GABA ligand 0.9 0.90.6 0.7 1.1 0.9 0.9 Cisapride 5-HT antagonist Peristaltic stimulant 1.00.9 0.7 0.8 1.1 0.9 0.8 Vigabatrin GABA transaminase inhibitorAnticonvulsant 0.9 1.0 0.6 0.8 1.0 0.9 0.9 Hydrastine hydrochlorideGABAa antagonist Hypotensor 1.0 0.9 0.6 0.8 0.8 0.8 0.9 Biperidenhydrochloride Anticholinergic Antiparkinsonian 1.0 0.9 0.6 0.8 1.0 1.01.0 Lobelanidine hydrochloride Nicotinic ligand — 0.9 0.9 0.7 0.7 1.10.9 0.9 Cetirizine dihydrochloride H1 antagonist Antihistaminic 0.9 1.00.6 0.8 0.8 1.0 0.9 Papaverine hydrochloride Phosphodiesterase inhibitorVasodilator 1.1 0.9 0.5 0.8 0.9 0.9 1.0 Etifenin Chelating agentDiagnostic agent 0.8 0.9 0.6 0.8 0.8 0.9 1.0 Yohimbine hydrochlorideAlpha antagonist Mydriatic 1.0 0.9 0.6 0.7 0.9 0.9 1.0 Metaproterenolsulfate, Beta-adrenergic agonist Bronchodilatator orciprenaline sulfate1.0 0.9 0.8 0.7 0.8 0.9 1.0 Lobeline alpha (−) hydrochoride Nicotinicreceptor ligand Respiratory stimulant 0.8 0.9 0.6 0.6 0.9 0.9 1.0Sisomicin sulfate Ribosomal protein synthesis Antibacterial inhibitor1.0 1.0 0.7 0.7 0.9 0.9 1.1 Berberine chloride Anti-HIV reverseAntibacterial transcriptase, cholinesterase inhibitor 0.7 0.8 0.1 0.60.4 0.6 0.9 Quercetine dihydrate Lipoxygenase inhibitor Antimalarial 0.90.9 0.8 0.7 0.9 0.9 0.9 Cilostazol Antithrombotic 0.0 0.0 0.0 0.0 0.00.0 0.1 Resveratrol Antiinflammatory 0.9 0.9 0.8 0.8 0.8 0.8 1.0Galanthamine hydrobromide Cholinesterase inhibitor, Alzheimer treatmentnicotinic receptor agonist 0.8 0.9 0.7 0.7 1.0 1.1 1.1 BromperidolDopamine antagonist Antipsychotic 0.8 0.9 0.6 0.8 0.8 0.9 0.9Bicuculline (+) GABAa receptor antagonist Convulsant 0.9 1.0 0.9 0.8 1.00.9 0.9 Cyclizine hydrochloride H1 antagonist Antiemetic 0.9 0.9 0.7 0.70.8 0.9 0.9 Yohimbinic acid monohydrate — — 0.9 0.9 0.7 0.6 0.8 0.8 1.0Chlorthalidone Na+ uptake inhibitor, Diuretic carbonic anhydraseinhibitor 0.9 0.9 0.4 0.6 0.8 0.8 1.1 Coralyne chloride hydrateTopoisomerase I inhibitor Anti-leukemic 0.8 0.8 0.1 0.8 0.7 0.6 1.1Dobutamine hydrochloride Beta1, Beta2 agonist Bronchodilatator 0.9 0.90.6 0.7 0.9 0.8 1.2 Corticosterone — Glucocorticoid 0.9 0.9 0.6 0.6 0.70.8 1.2 Moclobemide Mono amine oxidase Antidepressant inhibitor (Type A)0.8 0.8 0.5 0.6 0.8 0.7 1.2 Cyanocobalamin — Vitamin 0.8 0.9 0.7 0.7 0.80.8 1.0 Clopamide Gonad stimuling agent Antihypertensor 0.9 0.8 0.5 0.80.9 0.8 1.2 Cefadroxil Bacterial transpeptidase Antibacterial inhibitor0.9 1.2 0.8 0.7 0.8 0.8 1.1 Hycanthone DNA intercaling agentAnthelmintic 0.9 0.9 0.6 0.7 0.9 0.8 1.4 Cyclosporin A IL 2 synthesisinhibitor, Immunosuppressant calcineurine phosphatase inhibitor 0.8 0.80.6 0.6 0.7 0.8 1.2 Adenosine 5′-monophosphate Ca++ channel block,Nutrient monohydrate adenosine receptor activation, activation ofoutward K+ current 0.9 0.9 0.6 0.7 0.8 0.8 1.2 Digitoxigenin Na+ K+ATPase inhibitor Cardiotonic 10.6 1.4 0.4 0.7 1.7 0.7 1.0 AmoxicillinBacterial transpeptidase Antibacterial inhibitor 0.8 0.9 0.8 0.7 0.9 0.81.2 Digoxin Na+ K+ ATPase inhibitor Cardiotonic 0.8 0.9 0.7 0.7 0.9 0.81.4 Cephalexin monohydrate Bacterial transpeptidase Antibacterialinhibitor 1.0 1.7 0.9 0.7 1.0 0.6 1.2 Doxorubicin hydrochloride DNAintercalant Antibacterial 0.8 0.9 0.7 0.7 0.8 0.8 1.1 Dextromethorphanhydrobromide Opioid ligand Antitussive monohydrate 0.8 0.9 0.6 0.7 0.80.7 1.2 Carbimazole Iodine oxidazing inhibitor Antityroidic hormone 0.91.0 0.9 0.7 1.0 0.7 1.2 Droperidol Alpha adrenergic antagonist,Antipsychotic 5-HT antagonist, dopamine antagonist 0.9 1.0 0.6 0.7 0.80.8 1.1 Epiandrosterone — Anabolic steroid 1.0 0.9 1.1 0.8 1.0 0.9 1.1Fluoxetine hydrochloride 5-HT uptake inhibitor Antidepressant 1.0 0.90.7 0.7 0.9 0.9 1.3 Laudanosine (R,S) — Convulsant 1.0 0.9 0.7 0.7 0.90.8 1.1 Iohexol — Diagnostic aid 0.9 0.9 0.5 0.7 0.9 0.9 1.2 Ajmalicinehydrochloride Alpha antagonist Antihypertensor 6.3 0.8 0.6 0.9 1.1 1.01.3 Norcyclobenzaprine — Antiulcerative 0.9 0.9 0.8 0.7 0.9 0.8 1.2Trigonelline Antihyperglycaemic agent 0.9 0.8 0.7 0.8 0.9 0.8 1.2Pyrazinamide — Antibacterial 0.9 0.9 0.8 0.7 0.8 0.9 1.2 Diclofenacsodium Cyclooxygenase inhibitor Antiinflammatory 1.0 0.9 0.7 0.7 0.9 0.81.4 Trimethadione Ca++ channel blocker Anticonvulsant voltage dependant? 0.9 0.8 0.6 0.7 0.8 0.8 1.2 Calycanthine Calcium channel blocker 1.01.0 0.3 0.8 1.0 0.8 1.2 Lovastatin HMG CoA reductaseAntihypercholesterolemic inhibitor 1.1 0.9 0.6 0.7 1.0 0.8 1.3Convolamine hydrochloride Cholinergic ligand Vasodilatator 1.0 0.8 0.70.8 0.9 0.8 1.4 Nystatins Performs membrane Antifungal ionophores 0.90.9 0.7 0.7 0.9 0.8 1.3 Isocorydine (+) Ganglioblocker Hypotensor 0.90.9 0.6 0.7 0.9 0.9 1.4 Budesonide — Antiinflammatory 0.8 0.9 0.8 0.70.9 0.9 1.4 Xylazine Alpha-2 adrenergic agonist Antinociceptive 12.1 0.92.5 1.7 1.2 1.0 2.4 Imipenem Bacterial transpeptidase Antibacterialinhibitor 0.9 0.9 0.8 0.7 0.8 0.8 1.2 Seneciphylline — Antitumoractivity 0.8 0.9 0.8 0.7 1.1 0.8 1.1 Sulfasalazine15-hydroxydehydrogenase Treatment of ulcerative colitis inhibitor 0.90.9 0.8 0.7 0.8 0.9 1.2 Boldine Smooth muscle relaxant Choleretic 1.31.1 1.2 0.9 1.0 1.2 1.2 Bambuterol hydrochloride Beta2 adrenergicreceptor Bronchodilatator agonist 1.1 0.9 0.8 0.8 0.8 1.0 1.3Estradiol-17 beta — Estrogen 1.1 1.1 1.1 0.9 0.9 1.1 1.3 Betamethasone —Glucocorticoid 0.8 1.1 0.9 0.9 0.9 1.1 1.2 Fusaric acid Dopamine betahydroxylase Antibacterial inhibitor 1.2 1.0 1.0 0.9 1.0 1.0 1.1Colchicine Tubulin polymerisation Antiinflammatory inhibitor 1.0 1.1 1.10.8 1.1 1.0 1.3 Gabazine Antagonist GABA 1.2 1.0 1.1 1.0 1.3 1.1 1.3Metergoline 5-HT1 antagonist, D2 Antiprolactin agonist, 5-HT2 antagonist1.3 1.0 1.1 1.0 1.0 1.1 1.2 Ginkgolide A Cholinergic antagonistAlzheimer treatment 1.1 1.4 1.0 0.9 1.1 1.1 1.5 Brinzolamide Carbonicanhydrase inhibitor Antiglaucoma drug 1.3 1.0 1.0 0.9 1.2 1.1 1.2Cyclobenzaprine hydrochloride — Muscle relaxant 1.0 1.0 1.0 1.0 1.2 1.21.5 Ambroxol hydrochloride — Expectorant 1.2 1.0 1.1 0.8 1.0 1.1 1.1Carteolol hydrochloride Antihypertensor 1.4 0.9 1.2 1.1 1.0 1.1 1.4Benfluorex hydrochloride Increase of glucose Hypolipedimic penetrationand use by cells, decrease of triglyceride intestinal absorption 1.1 1.01.3 0.9 1.3 1.1 1.4 Hydrocortisone base — Glucocorticoid 1.3 1.1 1.6 1.01.5 1.2 1.4 Bepridil hydrochloride Ca++ channel blocker Antianginal 1.31.0 3.0 1.0 1.0 1.0 1.3 Hydroxytacrine maleate (R,S) Acetylcholineesterase inhibitor 1.0 0.9 2.0 0.8 0.9 0.9 1.2 Meloxicam Cyclooxygenaseinhibitor Anti-inflammatory 1.0 1.1 1.3 0.9 1.0 1.0 1.5 Pilocarpinenitrate Cholinergic Antiglaucoma drug 1.1 1.2 0.4 0.9 1.4 1.5 1.4Benzbromarone Uric acid transport inhibitor Coronarodilatator 1.2 1.07.1 0.9 1.5 1.4 1.2 Dicloxacillin sodium salt Bacterial transpeptidaseAntibacterial inhibitor 1.1 1.1 1.2 1.0 1.1 1.1 1.4 Glycocholic acidDetergent Anticholelithogenic 1.2 1.0 1.0 1.0 1.3 1.1 1.6 ScoulerineDopamine antagonist, Antiemetic alpha1 antagonist 1.3 1.0 0.4 1.0 1.10.9 1.4 Thiostrepton Antibacterial 1.2 1.1 1.0 0.9 1.1 1.1 1.6 AjmalineInhibitor of glucose uptake Antihypertensor by heart tissue 1.3 1.0 1.10.9 1.0 1.2 1.5 Methionine sulfoximine (L) Glutamine synthetase —inhibitor 1.3 1.1 1.0 1.0 1.2 1.2 1.3 Monocrotaline — For inducingpulmonary diseases in rats 0.6 0.5 0.6 0.4 0.5 0.6 1.3 TiabendazoleMicrotubule inhibitor Anthelmintic 1.2 1.0 1.0 1.0 1.0 1.1 1.6Piperlongumine — Antifungal 1.3 0.8 0.8 0.7 3.0 0.6 1.4 Rifampicin RNApolymerase inhibitor Antibacterial 1.2 1.0 1.0 0.9 1.3 1.2 1.4Hydrocotarnine hydrobromide — Hemostatic 1.1 1.0 1.3 1.0 1.2 1.1 1.4Ethionamide Antibacterial 1.3 1.0 0.9 0.8 1.1 1.1 1.6 (−)-CinchonidineAntimalarial 1.3 1.0 1.2 0.9 1.0 1.1 1.6 Tenoxicam Cyclooxygenaseinhibitor Antiinflammatory 1.2 1.1 1.0 1.0 1.1 1.3 1.4 Eburnamonine (−)— Cerebral vasodilatator 1.2 1.0 1.4 0.9 1.0 1.2 1.3 TriflusalCyclooxygenase inhibitor Antithrombotic 1.2 1.1 1.6 1.0 1.1 1.1 1.5Cinchonine Heme polymerase inhibitor Antimalarial 1.1 1.1 1.3 1.0 1.01.2 1.3 Mesoridazine besylate D antagonist Antipsychotic 1.2 1.1 1.6 0.91.1 1.1 1.4 Canavanine sulfate monohydrate Nitric oxide inductibleAnticancer agent (L,+) synthetase inhibitor 1.1 1.1 1.6 1.1 1.1 1.1 1.6Trolox — Vitamin E analog 1.1 1.0 1.4 0.9 1.0 1.0 1.3 Harmalinehydrochloride Monoamine oxydase Antihelminthic dihydrate inhibitor 1.31.0 1.1 0.9 1.1 1.1 1.7 Ketotifen fumarate H1 antagonist Antihistaminic1.0 1.0 1.0 0.9 1.1 1.1 1.4 Alizapride HCl Antiemetic 1.2 1.0 0.9 0.80.9 1.0 1.7 Debrisoquin sulfate Catecholamine depletor Antihypertensor1.0 1.0 0.8 0.9 1.1 1.0 1.2 Lactobionic acid — Antithrombonic 1.1 0.91.0 0.8 0.9 1.1 1.3 Amethopterin (R,S) Dehydrofolate reductaseAntineoplastic inhibitor 1.1 1.0 0.9 1.0 1.2 1.1 1.4 Lumicolchicinegamma Microtubule — depolymerizating agent 1.0 0.9 0.8 1.0 0.9 0.9 1.6Methylergometrine maleate 5-HT antagonist, alpha Oxytocic adrenergicagonist 1.1 0.9 0.9 0.8 1.0 1.1 1.6 Lysergol 5-HT antagonistAntipsychotic 1.4 1.0 1.3 1.3 1.5 1.1 1.8 Methiothepin maleate 5-HTautoreceptor — antagonist, 5-HT1c antagonist, 5-HT release inhibitorelectrical or K+ induced 1.1 1.0 0.9 0.8 0.9 1.1 1.5 Mebhydroline 1,5-H1 antagonist Antihistaminic naphtalenedisulfonate 1.2 1.0 4.6 1.1 5.01.0 1.8 Clofazimine — Antileprosy 0.8 1.0 4.3 0.3 3.2 0.5 2.1Meclocycline sulfosalicylate Ribosomal protein synthesis Antibacterialinhibitor 1.0 1.0 0.9 1.0 1.0 1.1 1.4 Nafronyl oxalate Phosphodiesteraseinhibitor Vasodilatator ?, 5-HT antagonist, bradykinine antagonist 1.21.2 2.1 0.9 1.9 1.2 1.4 Meclozine dihydrochloride Histamine antagonistAntiemetic 0.9 0.8 1.1 0.7 0.8 1.0 1.5 Bezafibrate Lipoprotein lipaseactivator Antihyperlipoproteinemic 0.9 1.0 1.3 0.8 1.2 1.1 1.4 MelatoninMelatonin receptor ligand Immunostimulant 1.0 1.0 1.4 0.9 0.9 1.1 1.5Mimosine Cell cycle blocker, apoptosis Anticancer agent inducer 1.0 1.11.4 1.0 1.0 1.2 1.5 Menadione — Vitamin K3 1.0 1.1 1.2 1.0 1.0 0.9 1.6Clebopride maleate — Spasmolytic 1.0 1.0 1.4 1.0 1.2 1.0 1.1 Dinoprosttrometamol Protaglandin agonist Smooth muscle activator 1.1 1.1 1.1 1.01.2 1.2 1.5 Pirenperone 5-HT2 antagonist 1.2 1.0 0.8 0.8 1.1 1.2 1.6Harmalol hydrochloride dihydrate — Vasorelaxant 1.3 0.9 1.0 1.0 1.0 1.21.5 Isoquinoline, 6,7-dimethoxy-1- — — methyl-1,2,3,4-tetrahydro,hydrochloride 1.4 1.0 0.9 0.9 1.1 1.1 1.6 Harmol hydrochloride Liverconjugation probe Anxiolytic monohydrate 1.2 0.9 0.8 0.9 1.1 0.9 1.4Phenacetin Cyclooxygenase inhibitor Analgesic 1.1 1.1 0.9 0.7 1.0 1.21.5 Harmine hydrochloride Topoisomerase II inhibitor Antibacterial 1.11.0 1.2 0.8 1.2 1.0 1.4 Atovaquone Antipneumocystic 1.3 1.3 0.8 1.0 4.11.2 1.8 Ellipticine Intercaling agent Anticancer 1.1 1.0 0.9 1.0 1.0 1.21.4 Methoxamine hydrochloride Alpha adrenergic agonist Antihypotensive1.1 0.9 0.1 0.8 0.8 1.0 1.4 Chrysene-1,4-quinone — — 1.2 1.0 1.2 0.9 1.11.1 1.7 (S)-(−)-Atenolol Blokage of the action of Antihypertensoradrenergic mediators on beta receptors 1.4 1.0 0.8 1.0 1.1 1.3 2.0Demecarium bromide Cholinergic (ophtalmic) 1.1 0.9 1.0 1.0 0.9 1.1 1.4Piracetam — Nootropic drug 1.1 1.0 1.0 0.8 1.0 1.2 1.3 Quipazinedimaleate salt 5-HT agonist, 5-HT3 ligand 1.2 0.9 1.1 1.1 1.1 0.9 1.3Phenindione Antivitamin K Anticoagulant 1.3 1.0 1.1 1.0 1.0 1.0 1.7Sparteine (−) Ganglioplegic Antiarrhythmic 1.0 1.1 1.2 0.8 0.9 1.0 1.7Thiocolchicoside GABA agonist ? Muscle relaxant 1.1 1.1 1.1 1.0 0.9 1.01.4 Diflorasone Diacetate Anti-inflammatory (topical) 1.0 1.4 1.5 1.01.0 1.1 1.7 Clorsulon Anthelmintic 1.1 1.0 1.3 0.6 1.0 0.9 1.4 Harmanehydrochloride Imidazoline receptors ligand Vasorelaxant 1.4 1.0 1.4 1.12.9 1.3 1.4 Lidoflazine Ca++ channel activator Coronary vasodilatator1.1 1.0 0.9 0.8 1.3 1.1 1.3 Tropisetron HCl 5 HT3 antagonist Antiemetic1.2 0.9 0.9 1.0 1.4 1.1 1.6 Betaxolol hydrochloride Beta adrenergicantagonist Antihypertensor 18.6 1.1 1.2 4.6 1.8 11.7 21.9 CefiximeAntibacterial 1.2 1.1 0.6 1.0 1.3 1.1 1.4 Nicardipine hydrochloride Ca2+channel antagonist Antihypertensor 1.2 1.0 1.2 1.0 1.3 1.2 1.3Metrizamide — Contrasting product 1.1 0.9 0.6 0.9 1.4 1.1 1.3 Probucol —Antihyperlipoproteinemic 1.1 1.0 1.0 1.0 1.5 1.1 1.4 Muramic acid,N-acetyl Constituent of bacterial peptidoglycan 1.5 0.8 0.7 1.0 1.5 0.91.2 Mitoxantrone dihydrochloride DNA topoisomerase II Antineoplasticinhibitor 0.9 0.8 0.5 0.9 0.8 0.9 1.2 Myricetin Neutral endopeptidase —inhibitor 1.4 1.3 1.5 1.0 2.2 1.2 1.5 GBR 12909 dihydrochloride Dopaminereuptake inhibitor Antidepressant 1.1 0.9 0.5 0.9 1.2 1.0 1.5Naringenine Antiestrogenic 1.4 1.0 1.1 1.0 1.4 1.2 1.4 Carbetapentanecitrate Sigma1 receptor ligand Antitussive 1.3 1.0 1.2 0.9 1.2 1.3 1.2Naringin hydrate — Anti-oxidant 3.0 1.1 1.2 1.6 3.2 2.3 1.8 Dequaliniumdichloride Blocker of the apamin- Antibacterial sensitive smallconductance Ca2+ activated K+ channel, detergent 1.3 1.0 1.2 1.0 1.3 1.11.2 Neostigmine bromide Cholinesterase inhibitor Spinal analgesic 1.30.9 1.2 1.2 1.5 1.1 1.5 Ketoconazole Cytochrome P450c17 Antifungalinhibitor, sterol 14- demethylase inhibitor 13.2 1.0 1.8 6.0 1.4 1.2 6.1Niridazole Helminthic DNA damage Anthelmintic 1.1 1.1 4.2 1.1 1.3 1.11.4 Fusidic acid sodium salt Protein synthesis inhibitor AntibacterialGTPase coupled 12.2 1.0 1.8 3.1 1.8 1.2 1.9 Ceforanide Antibacterial 1.11.6 0.7 0.9 1.2 1.0 1.2 Ciclopirox ethanolamine Cell membrane proteinsAntifungal synthesis inhibitor 1.4 1.0 0.8 1.0 1.3 1.2 1.4Methoxy-6-harmalan Benzodiazepine receptor Psoriasis treatment ligand1.4 1.0 1.0 1.1 1.3 1.2 1.4 Probenecid Uric acid uptake inhibitorUricosuric 1.4 1.0 1.0 1.1 1.4 1.2 1.5 Stachydrine hydrochloride Potentinhibitor of tyrosinase 1.4 0.9 0.9 1.1 1.2 1.1 1.3 Betahistine mesylateH1 agonist, H3 antagonist Vasodilatator 1.4 1.0 1.3 1.1 1.4 1.2 1.7Pyridoxine hydrochloride — Vitamin 7.5 0.9 1.1 2.0 5.7 17.1 1.5Tobramycin Ribosomal protein synthesis Antibacterial inhibitor 1.3 1.01.0 1.2 1.4 1.2 1.5 Cytisine (−) Partial nicotinic receptorAntiinflammatory antagonist 1.3 1.0 1.1 1.1 1.4 1.2 1.4 Tetramisolehydrochloride Alkaline phosphatase Antihelminthic inhibitor 1.4 0.9 1.01.2 1.2 1.1 1.5 Pseudopelletierine hydrochloride — — 1.4 1.1 1.1 1.1 1.31.1 1.5 Pregnenolone Estrogen 1.2 1.0 1.9 1.1 1.3 1.2 1.5 RacecadotrilEnkephalinase inhibitor Antidiarrhoeic 1.4 0.9 0.9 1.0 1.2 1.1 1.5Molsidomine NO° production ? Vasodilatator 1.3 1.1 1.0 1.0 1.3 1.1 1.3Folic acid — Vitamin Bc or M 1.2 0.9 1.1 1.1 1.3 1.1 1.6 Chloroquinediphosphate Heme polymerase inhibitor Antipaludic 1.2 1.0 1.1 1.2 1.21.1 1.5 Salsolinol hydrobromide Dopamine analog — 1.4 1.0 1.2 1.1 1.21.1 1.7 Trimetazidine dihydrochloride — Antianoxic 1.2 1.0 1.3 1.0 1.31.1 1.5 Gramine Cholinesterase inhibitor Antibacterial 1.5 1.0 1.3 1.11.3 1.2 1.4 Parthenolide MAP kinase inhibitor Antiinflammatory 1.3 1.01.3 1.2 1.5 1.2 1.5 Dimethisoquin hydrochloride Local anesthesic 1.2 1.01.1 1.3 1.1 1.1 1.4 Terbutaline hemisulfate Beta-2 adrenergic agonistBronchodilatator 1.2 1.0 1.1 0.9 1.2 1.1 1.7 Strophantine octahydrateNa+ K+ ion dependant Cardiotonic ATPase inhibitor 1.3 1.0 1.0 1.0 1.21.0 1.4 Ketanserin tartrate hydrate 5-HT1, 5-HT2, 5-HT2A Antihypertensorantagonist 1.1 1.1 1.1 1.0 1.1 1.0 1.4 Pantothenic acid calcium salt —Nutritional factor monohydrate 1.1 0.9 1.0 1.0 1.2 0.9 2.0 Hemicholiniumbromide Acetylcholine stores — depleter, acetylcholine uptake inhibitor22.1 1.2 2.5 8.4 2.0 1.2 25.3 Cefotetan Antibacterial 1.2 0.9 2.5 1.01.1 1.0 1.9 Kanamycin A sulfate Ribosomal protein synthesisAntibacterial inhibitor 0.4 0.3 0.2 0.3 0.4 0.4 1.6 Piperine Enzymeinhibitor Antidiarrhoeic 22.1 1.0 1.3 6.2 7.6 14.2 4.9 Amikacin hydrateRibosomal protein synthesis Antibacterial inhibitor 1.4 1.0 1.1 1.1 1.21.1 2.0 Brompheniramine maleate H1 Antagonist, Antihistaminicanticholinergic 1.2 1.1 1.4 1.0 1.2 1.0 1.4 Etoposide Topoisomerase IIinhibitor, Antineoplastic kinase inhibitor 1.2 0.9 2.1 1.0 1.1 1.0 1.4Primaquine diphosphate Heme polymerase inhibitor Antimalarial 1.8 4.42.0 1.6 5.4 1.4 1.9 Clomiphene citrate (Z,E) Antioestrogen, gonad-Ovulation inductor stimulating agent 1.0 1.1 1.1 1.1 1.1 1.0 1.7Progesterone — Progestogen 1.1 1.0 1.3 1.0 1.1 1.0 1.8 Oxantel pamoateFumarate reductase Anthelmintic inhibitor, neuromuscular depolarizingagent 1.2 1.3 2.5 1.0 3.0 1.0 1.7 Felodipine L-type Ca2+ channelsblocker Antihypertensor 1.2 1.1 1.1 1.2 1.6 1.2 2.0 Prochlorperazinedimaleate Dopamine antagonist Antiemetic 1.1 1.1 1.0 0.9 1.0 1.0 1.5Methoxy-8-psoralen Apoptosis inducer with UV Psoriasis treatment 1.1 1.11.3 1.0 1.2 1.0 1.4 Hesperidin — Anticancer 1.1 1.1 1.1 1.1 1.6 1.0 1.4Puromycin dihydrochloride Antimetabolite Antiprotozoal 1.5 1.1 1.2 1.43.0 1.1 1.6 Hexetidine Detergent Antifungal 1.4 1.0 1.1 1.4 1.3 1.3 1.3Thiamine hydrochloride — Vitamin 1.3 1.0 1.1 1.2 1.1 1.2 1.6 Selegilinehydrochloride Monoamine B oxydase Antiparkinsonian inhibitor 1.4 1.0 1.21.3 1.4 1.3 1.6 Dipivefrin hydrochloride Antiglaucoma drug 1.4 1.4 1.91.2 1.2 1.2 1.4 Pentamidine isethionate — Antiparasitic 1.2 1.0 1.3 1.31.2 1.2 1.5 Thiorphan Neutral endopeptidase Antinociceptive inhibitor1.5 1.0 1.0 1.4 1.2 1.4 1.6 Tolazamide ATP-sensitive K+ ion Antidiabeticchannels blocker 1.1 1.0 0.9 1.6 1.2 1.2 1.5 Riboflavine — Vitamin 1.21.0 0.7 1.8 1.1 1.2 1.5 Nifuroxazide Bacterial DNA damage Antibacterial1.2 1.1 1.0 1.3 1.2 1.2 1.6 Hydroquinine hydrobromide — Preventor ofmuscular cramp hydrate 1.2 1.1 1.1 1.5 1.2 1.2 1.3 Mycophenolic acidInosine monophosphate Antineoplastic dehydrogenase inhibitor 1.3 1.0 1.01.2 1.2 1.2 1.6 Epivincamine Antiaggregant 1.1 1.2 2.1 1.2 1.8 0.9 1.5Dirithromycin Ribosomal protein synthesis Antibacterial inhibitor 1.31.0 1.2 1.2 1.2 1.2 1.5 Retrorsine — Antineoplastic 1.2 1.0 1.0 1.2 1.31.2 1.8 Gliclazide K+ channel voltage Antidiabetic dependant inhibitor1.5 1.1 1.2 1.2 1.4 1.2 1.6 Conessine Antiamebic 1.4 1.0 1.1 1.3 1.2 1.31.7 DO 897/99 D3 antagonist — 1.1 1.0 1.2 1.2 1.2 1.1 1.3 ProtoveratrineA Acetylcholinesterase release Antihypertensor stimulant 1.6 1.2 1.6 1.31.9 1.3 1.6 Prenylamine lactate Ca++ channel activator Vasodilatator 1.21.0 1.2 1.2 1.3 1.2 1.7 Solanine alpha Butyrylcholinesterase Antifungalinhibitor 0.3 0.9 0.4 0.5 0.3 0.3 0.3 Sulmazole Phosphodiesterase IIICardiotonic inhibitor 0.4 1.0 0.4 0.4 0.4 0.4 0.3 Althiazide Na+ Cl−transport inhibitor Diuretic 0.4 0.9 0.3 0.4 0.3 0.3 0.3 Epicatechin-(−)— Antidiarrhoeic 0.4 0.8 0.4 0.4 0.3 0.4 0.3 Isopyrin hydrochlorideCyclooxygenase inhibitor Antipyretic 0.5 0.9 0.4 0.5 0.4 0.3 0.3Flunisolide — Glucocorticoid 0.5 1.0 0.6 0.5 0.8 0.4 0.3 Phenethicillinpotassium salt Bacterial transpeptidase Antibacterial inhibitor 0.5 0.80.4 0.4 0.4 0.3 0.7 N-Acetyl-DL-homocysteine Disulfure bridge breakerExpectorant Thiolactone 0.5 0.9 0.4 0.5 0.7 0.3 0.3Sulfamethoxypyridazine Inhibitor of folic acid Antibacterial synthesis0.5 0.8 0.4 0.5 0.5 0.4 0.3 Flurandrenolide Glucocorticoid 0.4 0.9 0.40.5 0.4 0.4 0.4 Deferoxamine mesylate Iron chelating agent 0.5 0.9 0.40.5 0.4 0.4 0.4 Helveticoside Inhibitor of membrane ATP Cardiotonic 0.50.8 0.4 0.5 0.4 0.4 0.4 Mephentermine hemisulfate Alpha-adrenergicreceptor Antihypotensive agonist 0.5 0.9 0.4 0.5 0.5 0.4 0.4 MyosmineCholinergic — 0.4 0.7 0.2 0.5 0.5 0.4 0.4 Ergocryptine-alphaVasoconstrictor 0.4 0.9 0.4 0.6 0.4 0.4 0.4 Betonicine — — 0.6 0.9 0.50.5 0.9 0.4 0.4 Sulfadimethoxine Inhibitor of folic acid Antibacterialbiosynthesis 0.5 0.8 0.4 0.5 0.5 0.4 0.4 Etanidazole DNA damageAntineoplastic adjunct (radiosensitizer) 0.5 0.9 0.4 0.6 1.5 0.5 0.4Sulfanilamide Inhibitor of folic acid Antibacterial biosynthesis 0.5 0.80.5 0.5 0.5 0.5 0.4 Butirosin disulfate salt Ribosomal protein synthesisAntibacterial inhibitor 0.5 0.9 0.4 0.7 0.5 0.5 0.4 Balsalazide SodiumAnti-inflammatory 0.7 0.8 0.6 0.6 0.7 0.6 0.7 Carbinoxamine maleate saltHistamine antagonist Antihistaminic 0.7 0.9 0.6 0.5 0.6 0.6 0.7 NiacinAntihyperlipoproteine-mic 0.5 1.5 0.6 0.5 0.5 0.6 0.7 MethazolamideAnhydrase carbonic inhibitor Diuretic 0.6 0.9 0.7 0.6 0.7 0.6 0.6Bemegride Respiratory stimulant 0.7 0.8 0.6 0.6 0.5 0.6 0.7Pyrithyldione — Sedative 0.7 0.8 0.6 0.6 0.6 0.5 0.7 Digoxigenin Mainlyused as a non- Diagnosis isotopic label for DNA 0.6 1.0 0.6 0.2 0.5 0.50.6 Spectinomycin dihydrochloride Ribosomal protein synthesisAntibacterial inhibitor 0.5 0.8 0.5 0.7 0.5 0.5 0.6 MeglumineExpectorant 0.6 0.9 0.7 0.6 0.5 0.5 0.6 Piromidic acid Topoisomerase IIinhibitor Antibacterial 0.6 0.8 0.5 0.6 0.5 0.5 0.6 CantharidinAphrodisiac 0.6 0.9 0.5 0.6 0.6 0.6 0.6 Trimipramine maleate saltnoradrednaline and 5-HT uptake inhibitor, alpha antagonist,anticholinergic, Dopamine and Histamine antagonist 0.5 0.9 0.2 0.6 1.10.4 0.5 Clioquinol Anti-infective 0.7 0.9 0.5 0.6 0.6 0.5 0.6Chloropyramine hydrochloride H1 antagonist Antihistaminic 0.5 0.7 0.40.4 0.4 0.5 0.6 Oxybenzone Ultraviolet screen 0.8 0.9 0.8 3.2 0.4 0.60.6 Furazolidone Monoamine oxidase Antiinfective inhibitor and protozoalDNA damage 0.6 0.8 0.5 0.5 0.6 0.6 0.5 Promethazine hydrochlorideAntihistaminic 0.6 1.1 0.5 0.5 0.5 0.5 0.5 Dichlorphenamide Carbonicanhydrase inhibitor 0.2 0.3 0.2 0.2 0.2 0.1 0.6 Chrysin 5-lipoxygenaseinhibitor Antifungal 0.8 3.5 0.7 0.9 2.6 0.5 0.6 Sulconazole nitrateErgosterol synthesis Antifungal inhibition 0.5 0.9 0.5 0.5 0.6 0.5 0.5Proxyphylline Vasodilator 0.5 1.4 0.5 0.7 0.7 0.7 0.9 GlimepirideAntidiabetic 0.6 0.9 0.7 0.6 1.0 0.6 0.8 Sulfaquinoxaline sodium saltInhibitor of folic acid Antibacterial synthesis 0.6 1.0 0.6 0.6 0.7 0.60.7 Picrotoxinin GABA channel blocker Analeptic 0.7 2.6 1.2 1.4 0.7 0.70.7 Streptozotocin Antineoplastic 0.6 1.0 0.6 0.6 0.7 0.6 0.7Mepenzolate bromide Anticholinergic and muscarinic antagonist 0.6 0.90.6 0.5 0.7 0.7 7.0 Metoprolol-(+,−) (+)-tartrate salt Beta adrenergicantagonist 0.7 0.9 0.5 0.6 0.7 0.6 0.8 Benfotiamine Vitamin 0.6 0.9 0.60.6 0.7 0.7 0.8 Flumethasone Antiinflammatory 0.6 0.9 0.6 0.5 0.7 0.70.8 Halcinonide Antiinflammatory 0.6 0.9 0.7 0.6 0.8 0.6 0.7 Flecainideacetate Antiarrhythmic 0.6 0.9 0.7 0.6 0.8 0.6 1.0 Lanatoside C Na+ K+ATPase inhibitor Cardiotonic 16.5 0.9 3.1 3.6 0.5 0.7 0.9 Cefazolinsodium salt Bacterial transpeptidase Antibacterial inhibitor 0.6 1.1 0.70.6 0.6 0.6 0.7 Benzamil hydrochloride Na+ channel blocker and blockerof Na+/Ca++ exchanger 0.7 0.9 0.7 0.6 0.8 0.6 0.8 Atractylosidepotassium salt Nucleotide transport Anticancer inhibitor 0.6 0.9 0.6 0.60.8 0.6 0.8 Suxibuzone — Antiinflammatory 0.7 1.0 0.7 0.6 0.7 0.6 0.7Folinic acid calcium salt — Antianemic 0.7 0.9 0.6 0.5 0.7 0.7 0.96-Furfurylaminopurine — Plant growth accelerator 0.7 1.0 0.6 0.8 0.8 0.70.7 Levonordefrin Adrenergic receptor agonist Vasoconstrictor 0.8 1.21.7 0.6 0.8 0.6 0.9 Avermectin B1 Ligand GABA receptors Antihelmetic 0.60.9 0.5 0.6 0.7 0.6 0.7 Ebselen Cyclooxygenase inhibitorAntiinflammatory 0.7 0.9 0.9 0.8 0.9 0.8 1.1 Bergenin monohydrate —Antiarrhythmic 0.7 0.9 0.6 0.6 0.8 0.8 1.1 3-Acetylcoumarin 0.7 0.9 0.70.7 0.9 0.8 1.0 Cromolyn disodium salt — Antiasthmatic 0.7 0.9 0.7 0.60.8 0.8 0.9 Esculin Hydrate Skin protectant 0.7 0.9 0.7 0.6 0.8 0.8 0.9Bucladesine sodium salt Adenylate cyclase modulator Cardiotonic 0.7 0.90.6 0.6 0.8 0.7 1.0 Felbinac Analgesic 0.9 0.9 1.0 0.7 1.1 8.6 0.9Cefsulodin sodium salt Bacterial transpeptidase Antibacterial inhibitor0.5 0.5 0.7 0.4 0.6 0.6 1.0 Butylparaben Antifungal 0.7 0.9 0.7 0.6 0.80.7 0.8 Fosfosal Cyclooxygenase inhibitor Analgesic 0.7 0.9 0.6 0.6 0.70.7 0.8 Aminohippuric acid Diagnostic aid (renal function) 0.7 0.9 0.60.7 0.8 0.7 0.9 Suprofen Cyclooxygenase inhibitor Analgesic 0.7 0.9 0.70.6 0.7 0.7 0.9 N-Acetyl-L-leucine Vertigo 0.7 0.9 0.7 0.6 0.7 0.6 1.0Catechin-(+,−) hydrate Antidiarrhoeic 14.7 1.1 0.7 1.7 0.9 0.9 1.0Pipemidic acid Antibacterial 0.8 0.9 0.7 0.6 0.7 0.6 0.8 NadololAdrenergic beta antagonist Antihypertensor 0.8 0.8 0.5 0.7 0.6 0.7 1.0Dioxybenzone Ultraviolet screen 18.1 1.3 4.4 3.9 0.6 0.6 0.8 Moxalactamdisodium salt Bacterial transpeptidase Antibacterial inhibitor 0.7 0.90.6 0.6 0.8 0.6 1.2 Adrenosterone Androgenic activity 0.6 0.9 0.6 0.60.7 0.8 0.8 Aminophylline Mastocytes degranulation Vasodilatatorinhibitor and benzodiazepines antagonist 0.7 1.0 0.6 0.6 0.7 0.7 0.8Methylatropine nitrate Mydriatic 0.8 1.0 0.7 0.8 0.7 0.8 0.8 VitexinAntiviral 0.9 0.9 0.6 0.7 0.8 0.8 0.9 Nadide Coenzyme nicotinamide —adenine dinucleotide 0.9 0.9 0.7 0.7 0.7 0.9 0.9 Gelsemine Tumourinhibitor 1.0 0.9 0.7 0.7 1.4 0.8 0.9 Sulfamethizole Inhibitor of folicacid Antibacterial synthesis 1.0 0.9 0.6 0.7 0.7 1.0 0.9 SolasodineCytotoxic 0.7 1.0 0.6 0.7 0.7 0.8 1.0 Medrysone — Glucocorticoid 0.8 0.90.7 0.7 0.9 0.9 1.2 Delcorine Nicotinic ligand Antiarrhythmic 0.6 0.60.5 0.5 0.5 0.6 1.0 Flunixin meglumine Analgesic 0.7 0.8 0.6 0.7 0.7 0.81.1 Evoxine Glycine receptor antagonist Sedative 0.9 1.0 6.6 0.8 0.9 0.71.2 Spiramycin Ribosomal protein synthesis Antibacterial inhibitor 0.90.9 0.4 0.7 1.4 0.8 1.0 Nisoldipine Antihypertensive 0.9 0.9 0.7 0.8 0.80.9 1.1 Glycopyrrolate Antispasmodic 0.9 0.9 0.7 0.8 0.8 0.8 1.0Foliosidine — Anticonvulsant 10.5 0.9 1.3 2.3 0.6 0.7 0.9 Cefamandolesodium salt Bacterial transpeptidase Antibacterial inhibitor 0.6 0.6 0.60.5 0.5 0.6 1.1 Skimmianine 5-HT ligand Sedative 0.9 1.1 0.7 0.6 0.7 0.91.1 Monensin sodium salt Performs membrane Antibacterial ionophores 1.00.9 0.6 0.8 0.8 0.9 1.0 Anabasine Insecticide 0.9 0.9 0.6 1.4 0.8 0.60.9 Isoetharine mesylate salt Beta adrenergic agonist Bronchodilator 0.91.0 0.9 0.9 0.9 0.9 1.1 Tetrandrine Analgesic 1.0 1.0 0.9 0.8 0.9 0.80.9 Mevalonic-D, L acid lactone HMG CoA substrate — 12.3 0.9 0.4 0.7 2.19.7 1.0 Azlocillin sodium salt Bacterial transpeptidase Antibacterialinhibitor 0.9 1.0 1.0 0.7 0.8 1.0 1.1 Hymecromone Antispasmodic 1.0 0.90.9 0.8 1.1 1.1 1.3 Clidinium bromide Anticholinergic Spasmolytic 1.00.9 0.8 0.8 0.8 0.9 1.0 Caffeic acid Antineoplastic 0.9 0.9 0.7 0.8 1.61.0 1.1 Sulfamonomethoxine Inhibitor of folic acid Antiseptic synthesis1.0 0.8 0.7 0.7 0.9 1.0 1.2 Diloxanide furoate Antiamebic 0.9 1.4 0.80.8 0.9 0.9 1.1 Benzthiazide Na+ Cl− transport inhibitor Diuretic 0.90.8 0.8 0.7 1.0 1.0 1.2 Metyrapone Diagnostic aid (pituitary function)0.8 1.4 1.3 0.8 1.0 0.8 1.2 Trichlormethiazide Na+ Cl− transportinhibitor Diuretic 1.0 0.9 0.9 0.7 1.0 1.0 1.1 Urapidil hydrochlorideAntihypertensor 0.9 0.8 0.8 0.7 0.8 0.9 1.2 Oxalamine citrate salt —Antiinflammatory 1.3 1.3 1.4 1.1 6.6 0.8 1.0 Fluspirilen Antipsychotic0.9 0.8 0.8 0.8 1.0 0.9 1.3 Propantheline bromide Muscarinic antagonistAntiulcerative 0.9 0.9 0.8 0.7 0.8 0.9 1.3 S-(+)-ibuprofen Analgesic 1.00.9 3.7 0.7 2.6 0.7 1.0 Lasalocid sodium salt Membrane ionophoresAntibacterial producer 0.8 0.9 0.5 0.7 0.9 0.9 1.1 Ethynodiol diacetateProgestogen 0.9 0.9 0.7 0.8 0.8 0.8 1.0 Dimethadione — Anticonvulsant0.5 0.4 0.4 0.5 0.4 0.6 1.0 Nabumetone Analgesic 0.9 1.0 0.5 0.7 0.9 0.91.0 Ethaverine hydrochloride Antispasmodic 0.9 1.0 0.7 0.7 0.8 0.9 0.9Nisoxetine hydrochloride Inhibitor of noradrenaline 0.9 1.0 0.7 0.8 0.80.9 1.2 Dydrogesterone Progestogen 0.8 0.8 0.8 0.7 0.7 0.6 1.2 Terazosinhydrochloride Alpha adrenergic antagonist Treatment of benign prostatichyperplasia 0.8 0.9 0.7 0.7 0.9 0.8 1.3 (d,l)-Tetrahydroberberine —Sedative 0.1 0.1 0.3 0.1 0.1 0.1 1.4 Phenazopyridine hydrochloride —Analgesic 0.9 0.9 0.8 0.7 0.8 0.8 1.1 Deltaline Nicotinic receptorligand Antiarrhythmic 0.5 0.9 7.1 0.2 3.4 0.4 1.3 Demeclocyclinehydrochloride Ribosomal protein synthesis Antibacterial inhibitor 0.80.8 0.7 0.6 0.8 0.8 1.2 Graveoline topoisomerase II inhibitor?Antihypotensive and CNS stimulant 0.8 0.9 0.8 0.7 0.8 0.8 1.4 Fenoprofencalcium salt dihydrate Cyclooxygenase inhibitor Antiinflammatory 0.9 0.90.7 0.6 0.8 0.9 1.4 Hippeastrine hydrobromide — Hypotensor 12.6 0.9 0.42.3 3.3 8.7 1.3 Piperacillin sodium salt Bacterial transpeptidaseAntibacterial inhibitor 0.9 0.9 0.8 0.7 0.8 0.7 1.4 Beta-EscinPeripheral vascular disorders 0.9 1.1 0.6 0.7 1.8 0.8 1.4Diethylstilbestrol Estrogen 0.6 1.3 0.2 0.3 0.4 0.5 1.5 Gossypol Ca2+uptake inhibitor Local contraceptive 0.9 1.0 0.8 0.8 0.8 0.8 1.4Chlorotrianisene Nuclear receptor ligand Non-steroidal estrogen 0.8 0.90.9 0.8 0.8 0.7 1.3 Ricinine 0.9 0.9 0.8 0.7 0.8 0.8 1.2 Ribostamycinsulfate salt Ribosomal protein synthesis Antibacterial inhibitor 0.9 0.90.8 0.7 0.9 0.8 1.3 Delsoline Nicotinic receptor antagonist andGanglioblocker 0.9 0.9 0.7 0.7 0.8 0.9 1.3 Methacholine chlorideCholinergic agonist 0.9 0.9 0.8 0.7 0.9 0.8 1.4 Fluorocurarine chlorideMuscle relaxant 0.9 0.9 0.8 0.7 0.7 0.8 0.9 Pipenzolate bromideNicotinic receptor antagonist Spasmolytic 0.9 0.9 0.8 0.8 0.9 1.0 1.4Butacaine Na+ channel blocker Local anesthesic 0.9 0.9 0.8 1.0 0.8 0.81.4 (+)-Isoproterenol (+)-bitartrate Sympathomimetic (VET) salt 20.1 1.03.4 3.2 1.3 0.8 1.2 Cefoxitin sodium salt Bacterial transpeptidaseAntibacterial inhibitor 0.7 0.6 0.6 0.5 0.5 0.7 1.4 MonobenzoneDepigmentor 9.0 0.8 0.8 0.7 0.8 0.9 1.4 Ifosfamide Antineoplastic 1.10.9 0.8 0.7 0.9 0.9 1.4 2-Aminobenzenesulfonamide Ligand and potentinhibitor of carbonic anhydrase B 3.2 1.5 4.5 0.7 1.1 1.0 1.6 Novobiocinsodium salt DNA topoisomerase IV Antibacterial inhibitor 0.8 0.8 0.7 0.70.8 0.8 1.4 Estrone Estrogen 0.9 0.9 0.8 0.8 1.1 0.9 1.3Tetrahydroxy-1,4-quinone — Keratolytic monohydrate 0.9 1.0 0.3 0.7 0.80.9 1.3 Lorglumide sodium salt CCKa ligand 0.2 0.1 0.2 0.1 0.2 0.2 1.5Indoprofen Cyclooxygenase inhibitor Analgesic 0.8 0.9 0.4 0.8 0.9 0.91.4 Nitrendipine Antihypertensor 0.8 0.9 1.6 0.7 1.1 0.9 1.4Carbenoxolone disodium salt Mucus stimulating synthesis Antiulcerative0.8 0.9 0.6 0.7 0.9 0.8 1.6 Flurbiprofen Anti-inflammatory 0.9 0.9 0.80.8 0.8 0.9 1.4 locetamic acid Diagnostic aid (radiopaque medium) 0.91.0 0.7 0.7 1.0 0.9 1.4 Nimodipine Vasodilator 0.9 0.9 0.9 0.7 0.8 0.91.4 Ganciclovir Antimetabolite Antiviral 1.0 1.3 0.4 0.8 1.1 0.8 1.2Bacitracin Antibacterial 0.9 0.9 0.7 0.8 1.0 1.0 1.3 Ethopropazinehydrochloride Anticholinergic Antiparkinsonian 0.8 0.9 0.8 0.7 0.8 0.81.2 L(−)-vesamicol hydrochloride Potent inhibitor of vesicularacetylcholine storage 1.2 1.1 1.2 1.0 1.0 1.1 1.3 Austricine hydrateAntiatherosclerotic 0.8 0.8 1.1 0.6 0.7 0.9 1.7 Butamben Na+ channelblocker Anesthetic 1.2 1.0 1.3 0.9 1.0 1.2 1.3 beta-BelladonnineMuscarinic receptor and dichloroethylate nicotinic receptor lignad 1.21.0 1.4 0.9 2.2 1.1 1.6 Sulfapyridine Antibacterial 1.1 1.0 1.1 1.0 1.11.1 1.5 Pempidine tartrate Nicotinic acetylcholine Antihypertensorreceptor antagonist 1.2 1.0 1.2 1.0 1.0 1.1 1.4 Meclofenoxatehydrochloride Acetylcholine precursor Cerebral stimulant 1.2 1.0 1.4 1.01.0 1.0 1.7 Heliotrine Model for hepatitis and cirrhosis in liver 0.91.1 0.7 2.6 1.0 1.1 1.7 Furaltadone hydrochloride Bacterial DNA damageAntibacterial 1.2 1.0 1.3 1.1 1.0 1.1 1.4 Nitrarine dihydrochloride —Hypotensor 1.1 0.9 1.1 0.9 0.9 1.1 1.4 Ethoxyquin — Antifungal 1.0 1.01.4 0.8 1.0 1.1 1.5 Lycorine hydrochloride Inhibitor of proteinAntitumoral translation 1.2 0.9 1.2 0.8 1.0 1.1 1.5 Tinidazole Protozoaland bacterial DNA Antiprotozoal damage 1.0 1.0 1.2 0.9 1.0 1.1 1.3Karakoline Inhibitor of ACE Hypotensor 1.3 1.0 1.2 0.7 1.0 1.1 1.4Guanadrel sulfate Antihypertensor 1.1 1.0 1.1 1.0 1.0 1.1 1.3Estropipate Menopause 1.1 1.1 1.5 0.7 1.0 1.0 1.8 Vidarabine Adenosineantimetabolite Antiviral 1.3 1.0 1.2 1.0 1.1 1.1 1.4 Ungerine nitrateEnhancer of analgesics Sedative 1.3 1.0 1.2 1.0 2.3 1.1 1.2 SulfameterInhibitor of folic acid Antibacterial synthesis 1.3 1.0 1.4 1.0 1.0 1.13.6 Napelline Anticholinergic agent Antiarrhythmic 1.3 1.1 1.2 1.1 1.01.2 1.5 Isopropamide iodide Anticholinergic 1.3 1.0 1.1 0.8 1.1 1.2 1.6Securinine — CNS stimulant 1.3 1.0 1.2 0.9 1.0 1.2 1.7 Nizatidine H2antagonist Antiulcerative 1.2 1.0 1.1 0.9 1.1 1.2 1.5 Trimeprazinetartrate Histamine antagonist Antipruritic 1.2 1.0 1.1 1.0 1.0 1.3 1.6Thioperamide maleate H3 antagonist Antiemetic 1.2 1.0 6.5 1.0 1.5 1.31.4 Nafcillin sodium salt Bacterial transpeptidase Antibacterialmonohydrate inhibitor 1.3 1.0 1.1 0.8 1.0 1.2 1.8 Xamoterol hemifumaratebeta1-Adrenoceptor Cardiotonic selective partial agonist 1.4 1.1 1.2 0.91.1 1.2 1.8 Procyclidine hydrochloride Muscarinic antagonistAntiparkinsonian 1.1 1.0 1.3 1.0 1.2 1.2 1.7 Rolipram Nootropic drug 1.31.1 1.3 0.9 1.2 1.1 1.5 Amiprilose hydrochloride — Immunomodulator 3.27.5 ### 4.7 5.6 1.5 1.8 Thonzonium bromide Detergent 1.2 0.9 0.8 0.8 1.11.1 1.5 Ethynylestradiol 3-methyl ether — Estrogen 1.1 1.0 1.1 1.0 1.11.2 1.5 Idazoxan hydrochloride Alpha2 agonist Antiparkinsonian 1.3 1.01.0 1.0 1.0 1.2 1.9 (−)-Levobunolol hydrochloride Adrenergic betaantagonist Antiglaucoma drug 1.2 1.1 1.2 1.0 1.2 1.2 1.6 Quinapril HClAngiotensin 1.3 1.1 1.3 1.0 1.0 1.2 1.5 Iodixanol Diagnostic aid(radiopaque medium) 1.2 1.0 1.3 1.0 1.2 1.2 1.4 NilutamideAntineoplastic 1.1 1.0 1.1 1.0 1.2 1.1 1.7 Rolitetracycline Ribosomalprotein synthesis Antibacterial inhibitor 1.3 1.0 1.1 1.0 1.0 1.0 1.4Ketorolac tromethamine Analgesic 1.3 1.0 3.9 0.9 1.0 1.1 1.4 Equilin —Estrogen 1.2 1.0 2.8 1.1 1.1 1.2 1.4 Protriptyline hydrochlorideAntidepressant 1.2 1.0 1.0 0.8 0.9 1.2 1.8 Fillalbin — — 1.1 1.0 1.1 1.10.9 1.0 1.2 Alclometasone dipropionate Corticoide Anti-inflammatory 1.01.0 1.0 0.9 0.9 1.2 1.5 Citalopram Hydrobromide 5HT uptake inhibitor 0.80.7 0.6 0.6 0.8 0.8 1.5 Leflunomide Inhibitor of T and B cellImmunosuppressive proliferation 1.2 1.0 1.4 0.8 0.9 1.1 1.7 Promazinehydrochloride Dopamine receptor Antipsychotic antagonist 1.0 1.0 1.1 1.11.1 1.1 1.7 Norgestrel-(−)-D Progestogen Oral contraceptive 1.1 1.0 0.91.0 2.1 1.0 1.9 Sulfamerazine Inhibitor of folic acid Antibacterialsynthesis 1.0 1.0 1.1 0.9 0.9 1.2 1.7 Fluocinonide Antiinflammatory 0.70.5 0.7 0.6 0.3 0.6 1.6 Acacetin Inhibitor of glutathione Antitumoragent reductase and of topoisomerase I 1.0 1.0 1.4 1.0 1.6 1.1 1.4Sulfamethazine sodium salt Inhibitor of folic acid Antibacterialsynthesis 1.2 1.0 1.2 1.0 1.1 0.9 1.6 Ethotoin Anticonvulsant 1.1 1.01.2 0.8 1.0 1.1 1.6 Guaifenesin — Expectorant 1.2 1.0 1.0 0.9 0.9 1.11.9 3-alpha-Hydroxy-5-beta- androstan-17-one 43.3 7.4 ### 23.4 12.3 19.21.5 Alexidine dihydrochloride Detergent Antibacterial 1.4 1.0 1.1 0.91.0 1.0 1.4 Tetrahydrozoline hydrochloride Adrenergic Vasoconstrictor1.2 1.1 0.8 0.8 1.1 1.1 1.7 Proadifen hydrochloride Cytochrome P450mono- Local anesthesic oxygenases inhibitor, Na+ channel blocker 1.4 1.20.7 0.7 3.0 1.1 1.6 Hexestrol Nuclear receptor ligand Estrogenantineoplastic 1.1 1.1 0.9 1.0 1.1 0.9 1.6 Zomepirac sodium saltCyclooxygenase inhibitor 20.0 1.4 5.0 4.6 1.4 1.0 1.6 Cefmetazole sodiumsalt Bacterial transpeptidase Antibacterial inhibitor 10.3 1.1 1.3 2.11.3 1.2 1.4 Cinoxacin Topoisomerase II inhibitor Antibacterial 1.4 0.91.4 1.2 1.3 1.3 1.6 Paroxetine Hydrochloride 5-HT uptake inhibitor 1.31.0 1.2 1.1 1.0 1.0 1.7 Propofol Anesthetic (intravenous) 2.0 1.1 9.40.3 3.4 0.7 1.5 Doxycycline hyclate Ribosomal protein synthesisAntibacterial inhibitor 0.9 1.0 1.6 1.1 1.0 1.3 1.8 S(−)Eticlopridehydrochloride 1.2 1.1 1.0 0.9 1.1 1.1 1.6 Liothyronine Thyroid hormoneThyroidic drug 1.0 0.9 1.3 0.9 1.2 1.2 1.8 Primidone Anticonvulsant 0.91.4 2.8 1.1 1.0 0.9 1.5 Roxithromycin Ribosomal protein synthesisAntibacterial inhibitor 1.1 0.9 1.1 0.8 1.2 1.1 1.8 FlucytosineAntifungal 1.1 1.0 0.5 0.9 1.0 1.0 1.5 Beclomethasone dipropionate —Antiinflammatory 1.2 1.0 1.1 1.0 1.1 1.3 1.9 (−)-MK 801 hydrogen maleateNMDA antagonist Anticonvulsant 0.9 0.8 0.9 0.6 0.8 1.0 2.0 Tolmetinsodium salt dihydrate Cyclooxygenase inhibitor Antiinflammatory 1.2 1.01.2 1.0 0.9 1.0 2.0 Bephenium hydroxynaphthoate Antihelmintic 1.2 0.91.3 0.9 0.9 1.2 1.5 (+)-Levobunolol hydrochloride Beta adrenergicantagonist Antiglaucoma drug 1.0 1.1 0.8 0.9 1.1 1.1 1.7Dehydroisoandosterone 3-acetate Menopausal syndrome 1.4 1.1 1.2 0.9 1.31.1 1.5 Doxazosin mesylate Alpha 1 adrenergic Antihypertensor antagonist1.3 0.9 1.4 1.2 1.0 1.1 1.8 Benserazide hydrochloride Inhibitor ofL-aromatic In combination with levodopa amino acid decarboxylase asantiparkinsonian 1.3 1.0 1.0 0.9 0.9 1.0 1.7 Fluvastatin sodium salt HMGCoA reductase Anti-hyperlipoproteinemic inhibitor 1.2 1.0 1.0 1.0 1.21.1 1.6 Iodipamide Diagnostic aid (radiopaque medium-cholecystographic)1.4 1.1 1.3 1.0 1.0 1.1 2.0 Methylhydantoin-5-(L) — — 1.4 1.0 1.1 1.21.1 0.9 1.4 Esculetin Antifungal 1.2 1.1 1.4 1.1 1.4 1.3 1.5Trihexyphenidyl-D,L Anticholinergic Antiparkinsonian Hydrochloride 1.21.0 0.9 1.0 1.3 1.1 1.3 Clobetasol propionate — Glucocorticoid 1.4 1.01.3 1.1 1.3 1.1 1.4 Succinylsulfathiazole Inhibitor of folic acidAntibacterial synthesis 1.2 0.9 1.0 1.2 1.2 1.1 1.3 PodophyllotoxinAntiviral 1.2 1.0 0.8 1.1 1.1 1.2 1.5 Famprofazone Antipyretic 1.2 1.11.4 1.1 1.3 1.2 1.5 Clofibric acid Lipoproteine lipase activator ?Antihyperlipoproteinemic 1.3 0.9 1.3 1.0 1.3 1.1 1.3 Bromopride Dopamineantagonist Antiemetic 1.3 1.2 1.3 1.1 1.3 1.1 1.5 BendroflumethiazideNa+ Cl− transport inhibitor Diuretic 5.1 7.4 ### 7.4 5.8 8.9 1.3 Methylbenzethonium chloride Detergent Antibacterial 1.1 1.3 1.3 1.1 1.3 1.21.5 Dicumarol Vitamin K antagonist Anticoagulant 1.3 1.0 1.2 1.1 1.5 1.11.6 Chlorcyclizine hydrochloride Histamine antagonist Antihistaminic 1.20.9 1.0 1.0 1.4 1.0 27.1 Methimazole Iodine oxidazing inhibitorAntihyperthyroid 1.3 1.1 1.2 1.0 1.3 1.1 1.4 Diphenylpyralinehydrochloride H1 antagonist Antihistaminic 0.5 0.6 0.2 0.4 0.7 0.4 1.4Merbromin — Antibacterial 2.9 6.2 ### 5.7 5.4 5.8 1.3 Benzethoniumchloride Detergent Antibacterial 1.2 1.0 1.3 1.0 1.3 1.1 1.4 Hexylcainehydrochloride Local anesthesic 1.2 0.9 0.9 1.0 1.2 1.0 1.3 Trioxsalen —Pigmentation agent (photosensitizer) 1.5 1.0 1.5 1.2 1.3 1.2 1.5Drofenine hydrochloride — Spasmolytic 1.4 1.0 1.3 1.1 1.3 1.1 1.3Strophanthidin ATPase inhibitor Cardiotonic 1.3 1.0 1.1 1.0 1.2 1.1 0.9Cycloheximide — Antibacterial 1.4 1.0 1.5 1.1 1.4 1.3 1.6 GabapentinGABA receptor ligand Anticonvulsant 1.3 1.0 1.4 1.1 1.3 1.2 1.5 Penteticacid Chelating agent (iron) 1.5 0.9 1.2 1.1 1.5 1.2 1.3 Raloxifenehydrochloride 1.3 1.0 1.3 1.1 1.3 1.2 1.6 Bretylium tosylateAntiadrenergic 1.3 1.0 1.2 1.0 1.2 1.1 1.4 Etidronic acid, disodium salt— Calcium regulator 1.3 1.0 1.3 1.0 1.4 1.2 1.5 Pralidoxime chloride 1.21.0 1.2 1.1 1.3 1.2 1.5 Methylhydantoin-5-(D) — — 1.4 0.9 1.1 1.2 1.31.1 1.4 Phenoxybenzamine hydrochloride Nonselective alpha-Antihypertensor adrenergic blockade 1.6 1.0 0.4 1.2 2.5 1.1 1.5Simvastatin HMG-CoA reductase Antihyperlipidemic inhibitor 1.4 1.0 1.91.1 1.5 1.2 1.5 Salmeterol beta 2 adrenergic Bronchodilator agonist 2.21.0 1.2 0.9 1.4 1.2 1.5 Azacytidine-5 Antimetabolite Antineoplastic 1.31.0 1.1 1.0 1.3 1.2 1.4 Altretamine 1.4 0.9 1.3 1.0 1.5 1.0 1.4Paromomycin sulfate Ribosomal protein synthesis Antiamebic inhibitor 1.21.1 1.4 0.9 1.4 1.1 1.5 Prazosin hydrochloride Antihypertensor 1.4 1.01.6 1.2 1.5 1.1 1.6 Acetaminophen Cyclooxygenase inhibitor Antipyretic1.3 1.0 1.3 1.1 1.5 1.1 1.4 Timolol maleate salt Antiglaucoma drug 1.41.0 1.4 1.0 1.4 1.0 1.2 Phthalylsulfathiazole Inhibitor of folic acidAntibacterial synthesis 1.3 1.0 1.3 1.1 1.3 1.0 1.4 (+,−)-Octopaminehydrochloride beta-3 Adrenergic receptor Adrenergic agonist 0.4 0.5 0.20.5 0.3 0.5 1.6 Luteolin — Expectorant 1.4 1.0 1.3 1.2 1.3 1.0 1.3(±)-Nipecotic acid Activates GABA-like ion channels 1.1 1.1 1.2 1.1 2.81.1 1.6 Sulfabenzamide Inhibitor of folic acid Antibacterial synthesis1.1 1.0 1.0 1.0 1.1 1.0 1.7 (R)-Naproxen sodium salt Cyclooxygenaseinhibitor Antiinflammatory 1.1 1.0 1.2 0.9 1.2 1.1 1.6 Benzocaine Na+channel blocker Anesthetic 1.3 0.8 1.4 0.8 1.1 1.0 1.6 Propidium iodideDetergent Antibacterial 1.1 1.0 1.4 1.1 1.1 1.1 1.4 DipyroneCyclooxygenase inhibitor Antipyretic 1.2 1.1 1.3 1.2 1.4 1.2 2.0Cloperastine hydrochloride Histamine antagonist Antitussive 1.1 1.0 1.21.0 1.1 1.0 1.8 Isosorbide dinitrate Nitric oxide (NO) donor Antianginal1.2 1.1 1.3 0.9 1.1 1.1 1.6 Eucatropine hydrochloride Anticholinergic1.0 1.0 1.1 1.1 2.1 1.0 1.5 Sulfachloropyridazine Inhibitor of folicacid Antibacterial synthesis 1.1 1.0 1.5 1.1 1.1 1.1 1.5 IsocarboxazidAntidepressant 1.1 1.0 1.3 1.1 1.1 1.2 1.5 Pramoxine hydrochloride Localanesthesic 0.8 1.2 1.1 0.8 1.0 1.0 1.7 Lithocholic acidAnticholelithogenic, Cholagogue gastrointestinal agent 1.1 1.1 1.0 0.91.2 1.1 1.8 Finasteride Antialopecia agent 1.3 1.0 1.2 1.0 1.1 1.1 1.8Methotrimeprazine maleat salt — Analgesic 1.1 1.0 1.2 1.0 1.3 1.1 1.7Fluorometholone — Glucocorticoid 1.1 1.3 0.4 0.9 3.2 1.1 21.9 DienestrolNuclear receptor ligand Non-steroidal estrogen 11.8 1.0 5.3 3.2 1.1 1.12.0 Cephalothin sodium salt Bacterial transpeptidase Antibacterialinhibitor 1.1 1.1 1.2 0.9 1.1 1.1 1.6 Pridinol methanesulfonate saltAnticholinergic Antiparkinsonian 12.1 1.0 3.4 2.4 1.4 1.6 1.6 Cefuroximesodium salt Bacterial transpeptidase Antibacterial inhibitor 1.3 1.0 1.11.1 1.2 1.0 44.0 Amrinone TNF production inhibitor 1.2 1.0 1.3 1.2 1.21.2 1.4 Iopamidol Diagnostic aid (radiopaque medium) 1.5 1.1 1.3 1.1 1.31.3 1.3 Crotamiton Scabicide 1.2 1.0 1.3 1.1 1.1 1.2 1.4 IopromideContrast molecule 1.2 1.1 1.4 1.1 1.2 1.2 1.4 Propranolol hydrochlorideBeta blocking agent Antiarrhythmic 1.2 1.0 1.3 1.1 1.2 1.2 1.3Theophylline monohydrate Mastocyte degranulation Bronchodilatatorinhibitor in vitro 1.3 1.0 1.2 1.2 1.3 1.2 1.3 (R)-(+)-Atenolol 1.2 1.11.3 1.2 1.1 1.2 1.4 Theobromine Adenosine receptor Cardiotonicantagonist 1.4 1.0 1.4 1.5 1.3 1.2 1.2 Tyloxapol Mucolytic 1.3 1.1 1.31.2 1.5 1.4 1.5 Reserpine 1.2 1.0 ### 0.3 1.2 1.4 1.4 FlorfenicolAntibacterial 0.7 0.6 0.9 0.7 0.8 0.8 1.2 Arcaine sulfate Lowers bloodsugar 1.1 1.2 0.9 1.0 1.2 1.2 1.6 Megestrol acetate Progestogen 1.2 1.01.2 1.1 1.2 1.1 1.5 Scopolamine hydrochloride Non-selective muscarinicAntiemetic antagonist 1.2 1.2 1.6 1.2 1.3 1.3 1.3 DeoxycorticosteroneCorticoide Antiinflammatory 1.2 1.0 1.2 1.2 1.3 1.2 1.4 IoversolDiagnostic aid (radiopaque medium) 1.2 1.0 1.2 1.0 1.3 1.2 1.4 UrosiolAnticholelithogenic 1.3 1.1 1.2 1.0 1.2 1.1 1.5 Capsaicin Vanilloidreceptor ligand Topical analgesic 1.3 1.0 1.2 1.1 1.4 1.1 1.2Proparacaine hydrochloride Local anesthesic (VET) 1.2 1.1 1.4 1.2 1.31.2 1.2 Carbachol Cholinergic agonist, Myotic Cholinesterase inhibitor ?1.2 1.1 1.2 1.2 1.3 1.1 1.6 Aminocaproic acid Antiallergic 0.5 0.9 0.40.4 0.3 0.4 0.4 Denatonium benzoate Bittering agent to prevent poisoning0.5 0.9 0.3 0.4 0.3 0.4 0.3 Etomidate Hypnotic 0.4 0.9 0.3 0.4 0.4 0.40.3 Scopoletin Eicosanoid release inhibitor Antispasmodic 0.4 0.8 0.40.4 0.4 0.4 0.3 Tridihexethyl chloride Antispasmodic 0.5 1.1 0.4 0.5 0.40.4 0.4 Enilconazole Antifungal 0.5 0.8 0.4 0.5 0.4 0.5 0.4 Penbutololsulfate Antiarrhythmic 3.2 0.9 1.9 0.2 4.2 0.2 0.4 Methacyclinehydrochloride Antibacterial 0.4 0.9 0.4 0.5 0.4 0.4 0.3 PrednicarbateGlucocorticoid (topical) 0.5 0.8 0.4 0.5 0.4 0.4 0.4 Gibberellic acidPlant growth regulator 0.7 2.8 0.7 0.7 2.4 0.5 0.4 Sertaconazole nitrateAntibacterial 0.4 0.8 0.4 0.5 0.5 0.4 0.4 Sotalol hydrochlorideBeta-blocker 0.4 0.9 0.3 0.4 0.4 0.4 0.4 Repaglinide Antidiabetic 0.50.9 0.4 0.5 0.5 0.4 0.4 6-Hydroxytropinone 0.5 0.8 0.4 0.5 0.4 0.4 0.4Piretanide Antihypertensor 0.5 0.9 0.4 0.5 0.5 0.4 0.5 Decamethoniumbromide Muscle relaxant (skeletal) 0.5 0.9 0.5 0.5 0.6 0.5 0.4Piperacetazine Antipsychotic 0.5 0.8 0.4 0.4 0.4 0.4 0.43-Acetamidocoumarin Antiinflammatory 0.5 0.9 0.4 0.5 0.5 0.5 0.5Oxyphenbutazone Inhibition of cyclo-oxygenase Anti-inflammatory 0.4 0.80.6 0.4 0.5 0.5 0.5 Roxarsone Control of enteric infections. To improvegrowth and feed efficiency (VET) 0.6 0.9 0.4 0.6 0.5 0.5 0.4Quinethazone Diuretic 0.7 0.9 0.6 0.6 0.7 0.7 0.8 RemoxiprideHydrochloride Dopaminergic antagonist Antipsychotic 0.6 1.0 0.6 0.6 0.60.6 0.7 Moricizine hydrochloride Antiarrhythmic 0.6 1.0 0.6 0.6 0.6 0.60.8 THIP Hydrochloride GABAergic Agonist 0.6 0.9 0.4 0.6 0.6 0.6 9.6Iopanoic acid Contrast molecule 0.7 0.9 0.6 0.6 0.7 0.7 0.7 Pirlindolemesylate Highly selective reversible Anti depressant inhibitor ofmonoamine Seizures oxidase type A 7.7 0.9 0.6 0.5 1.1 0.6 0.8Pivmecillinam hydrochloride Antibacterial 0.7 0.9 0.6 0.6 0.6 0.6 0.9Pronethalol hydrochloride Antihypertensor 0.6 1.0 0.6 0.6 0.7 0.7 0.7Levopropoxyphene napsylate Antitussive 0.9 0.8 0.5 0.6 0.7 0.7 0.9Naftopidil dihydrochloride Alpha1 agonist Antihypertensor 0.7 1.0 0.60.6 0.6 0.7 0.7 Piperidolate hydrochloride Antispasmodic 0.5 1.1 0.5 0.60.8 0.6 0.7 Tracazolate hydrochloride GABA receptor ligand 0.9 0.9 0.71.1 1.1 0.6 0.7 Trifluridine Antiviral (ophthalmic) 0.6 1.0 0.6 0.7 0.70.7 0.9 Zardaverine Phosphodiesterase III & IV inhibitor 0.7 0.9 0.6 0.60.6 0.6 0.9 Oxprenolol hydrochloride Blocking beta-adrenergicAntiarrhythmic receptors 0.6 0.9 0.6 0.5 0.7 0.6 0.7 MemantineHydrochloride NMDA receptor antagonist Altzheimer disease 0.7 1.1 0.60.6 0.7 0.7 0.8 Ondansetron Hydrochloride Antagonist of 5- Antiemetichydroxytryptamine (serotonin) subtype 3 (5-HT 3) receptors 0.6 0.9 0.50.5 0.7 0.6 0.9 Ozagrel hydrochloride Thromboxane synthase Antianginalinhibitor 0.6 1.0 0.7 0.6 0.8 0.7 0.8 Propoxycaine hydrochloride Localanesthesic 0.6 0.9 0.6 0.5 0.7 0.6 1.0 Piribedil hydrochlorideDopaminergic agonist Vasodilator (peripheral) 0.6 1.1 0.5 0.6 0.7 0.70.9 Oxaprozin Analgesic 0.9 0.9 0.6 0.7 0.7 0.8 1.2 Nitrocaramiphenhydrochloride Muscarinic antagonist 1.0 0.9 0.7 0.8 0.8 0.7 1.1Phensuximide Anticonvulsant 0.9 1.0 0.7 0.7 0.8 0.9 1.0 NandroloneAnabolic 0.9 0.9 0.6 0.8 0.8 0.8 1.1 loxaglic acid Low-osmolar, ioniccontrast Diagnostic aid (radiopaque medium medium) 0.8 0.9 0.7 0.7 0.80.8 1.0 Dimaprit dihydrochloride Histamine H2 receptor agonist 1.0 1.00.8 0.7 1.1 0.8 1.0 Naftifine hydrochloride Antifungal 0.9 0.9 0.6 0.70.8 0.8 1.0 Reserpinic acid hydrochloride 0.9 0.9 0.6 0.7 0.8 0.9 0.8Meprylcaine hydrochloride Local anesthesic 0.8 0.9 0.7 0.8 0.8 0.8 1.1Beta-sistosterol Anticholesteremic 0.8 0.9 0.7 0.8 0.8 0.9 0.9 MilrinoneCardiotonic 0.9 0.8 0.6 0.7 0.8 0.9 0.9 Proscillaridin A Glucosidecardiotonic 0.9 0.9 0.7 0.8 0.8 0.9 1.1 Methantheline bromideAntispasmodic 0.8 0.9 0.7 0.8 0.8 0.7 1.2 CiprofibrateAntihyperlipoproteine-mic 0.8 0.9 0.7 0.7 0.8 0.8 1.5 Fluticasonepropionate Anti-inflammatory 0.9 0.9 0.7 0.8 0.8 0.8 1.2 Tropine 1.2 1.00.7 1.0 1.9 0.9 1.3 Zuclopenthixol hydrochloride Blocking of D1 and D2Schizophrenia dopaminergic receptors 11.3 1.7 0.8 0.6 2.9 0.7 1.2Benzylpenicillin sodium Antibacterial 0.9 0.8 0.9 0.7 0.9 0.8 1.1Proguanil hydrochloride Inhibition of dihydrofolate reductase 0.9 0.90.8 0.7 0.9 0.8 1.2 Chlorambucil Antineoplastic 0.8 1.0 0.7 0.7 0.8 0.71.2 Lymecycline Antibiotic 0.4 0.5 1.3 0.4 0.5 0.5 1.3 Methiazole 1.21.1 1.0 0.9 1.0 1.0 1.4 Alfadolone acetate In combination withalfaxalone as anesthesic (intravenous) 1.2 1.1 1.0 1.0 1.0 1.1 1.6(S)-propranolol hydrochloride Beta-adrenergic blocking Antihypertensoragent 1.2 1.1 1.2 1.0 1.0 1.2 1.7 Alfaxalone In combination withalfadolone acetate as anesthetic (intravenous) 1.0 1.1 0.9 0.9 0.9 0.81.4 (−)-Eseroline fumarate salt Anti-acetylcholinesterase Potentanalgesic activity & opiate agonist activity 1.1 1.0 0.8 0.9 0.9 1.0 1.5Azapropazone Cyclooxygenase inhibitor Analgesic 1.1 1.1 1.1 1.0 1.1 1.01.2 Condelphine 1.3 1.1 0.9 0.9 1.1 1.1 1.4 Meptazinol hydrochlorideAnalgesic (narcotic) 1.3 1.1 0.9 1.0 1.0 1.1 1.6 Leucomisine 1.2 1.0 0.90.8 1.0 1.1 1.4 Apramycin Antibacterial 1.4 1.0 2.4 0.9 1.1 1.0 1.3Dubinidine 1.2 1.0 0.9 0.9 1.1 1.1 1.3 Epitiostanol Antineoplastic 1.41.2 1.4 1.1 1.0 1.2 1.2 D-cycloserine Competitive inhibition of alaninracenase and D-alanin synthase (bacterial cell wall) 1.4 1.1 1.0 1.0 1.31.1 1.2 Fursultiamine Hydrochloride Altzheimer's disease 1.0 1.1 1.1 0.91.2 1.0 1.4 2-Chloropyrazine Cardiotonic 1.1 1.1 1.3 0.9 1.1 1.1 1.4Gabexate mesilate Anticoagulant 1.3 1.1 1.0 1.0 1.1 1.1 1.3(+,−)-Synephrine Hypertensive 1.5 1.1 0.4 0.9 1.8 0.9 1.2 PivampicillinAntibacterial 1.3 1.1 1.1 1.0 1.1 1.1 1.6 (S)-(−)-CycloserineTuberculosis 15.2 1.4 1.2 0.8 0.8 1.1 1.6 Talampicillin hydrochlorideAntibacterial 1.3 1.1 1.2 1.1 1.8 1.1 1.5 Homosalate Tool for uv screen1.3 1.0 6.1 1.1 1.5 1.2 1.3 Flucloxacillin sodium Antibacterial 1.4 1.11.0 0.9 0.9 1.1 1.6 Spaglumic acid NMDA receptor ligands 1.3 1.1 0.9 0.91.0 1.0 1.5 Trapidil Vasodilator (coronary) 1.2 0.9 1.0 0.9 1.2 1.0 1.4Ranolazine Antianginal 1.5 1.1 1.1 0.9 1.3 1.0 1.6 Deptropine citrateChronic bronchitis 1.1 1.0 1.2 1.0 1.0 0.9 1.7 (−)-Quinpirolehydrochloride D2-dopamine receptor agonist, some selectivity for D3sites 1.6 1.1 1.6 1.4 2.9 1.2 1.5 Sertraline 5-HT uptake inhibitor 1.01.0 0.9 1.0 2.9 1.0 1.3 Sulfadoxine Inhibition of Antibacterialdihydropteroate synthase 1.2 0.9 1.0 1.1 1.2 1.0 0.8 EthamsylateRetinopathy 1.2 1.0 1.1 1.0 1.0 1.1 1.3 Cyclopentolate hydrochlorideMydriatic 1.1 0.9 1.0 1.1 1.1 1.1 1.8 Moxonidine Imidazoline receptorligand Antihypertensor 1.1 1.1 1.2 0.8 1.2 1.0 1.4 Estriol Estrogen 1.01.0 1.1 0.8 1.4 1.1 1.5 Etilefrine hydrochloride Adrenergic agonistAntihypotensive 1.2 0.9 0.9 1.2 0.9 1.1 2.3 (−)-Isoproterenolhydrochloride Beta Adrenergic Agonist Cardiovascular drug (Sick SinusSyndrome) 1.3 1.0 1.1 1.1 1.0 1.1 1.4 Alprostadil Vasodilator 0.5 1.00.2 0.6 0.5 0.6 1.6 Kaempferol Topoisomerase I inhibitor,Anti-inflammatory tyrosin kinase, xanthin oxidase 0.9 1.1 0.6 0.9 1.71.1 1.5 Tribenoside Antihaemorrhoidic drug 1.2 1.1 1.2 0.9 1.0 1.2 1.4Nialamide Antidepressant 0.9 0.9 1.1 0.8 0.9 1.0 4.9 RimexoloneCorticoid Anti-inflammatory (local) 0.9 1.0 1.8 0.8 1.1 1.1 1.6 VitaminK2 Dietary factor for controlling blood pressure 1.1 1.1 0.7 1.1 1.2 1.11.4 Isradipine Antianginal 1.2 1.0 1.1 1.1 1.4 1.2 1.3 PerindoprilAntihypertensive 1.2 1.0 1.1 1.1 1.6 1.1 1.4 Tiletamine hydrochlorideAnesthetic 1.2 1.0 1.1 1.1 1.3 1.0 1.3 Fexofenadine HCl Antihistaminic1.2 1.0 1.2 1.1 1.5 1.1 1.2 Isometheptene mutate AdrenergicSympathomimetic (VET) 1.2 1.0 1.2 1.3 1.4 1.1 1.2 Quinic acid 1.1 1.00.8 1.2 1.5 1.1 1.3 Nifurtimox Antiprotozoal (Trypanosoma) 1.0 0.9 1.21.1 1.4 1.0 1.5 Clonixin Lysinate Analgesic 1.2 1.0 1.2 1.1 1.3 1.0 1.4Letrozole Antineoplastic 0.7 1.2 0.5 0.8 0.9 0.8 1.6 VerteporfinTreatment of age-related macular degeneration 1.2 0.9 1.1 1.2 1.5 1.11.4 Arbutin Tyrosinase inhibitor and Antibacterial Melanin biosynthesisinhibitor 13.4 1.0 3.9 3.6 1.5 14.8 1.2 Meropenem Antibacterial 1.2 1.01.1 1.1 1.6 1.1 1.4 Tocainide hydrochloride Antiarrhythmic 1.1 1.0 1.21.0 1.3 1.2 1.1 Ramipril Converting enzyme inhibitor Antihypertensor15.6 1.6 1.0 1.0 5.9 1.1 1.6 Benzathine benzylpenicillin Antibacterial1.1 1.0 1.1 1.0 1.3 1.0 1.3 Mephenytoin Anticonvulsant 1.1 1.0 1.3 1.11.5 1.1 1.3 Risperidone 5-HT2 antagonist Antipsychotic 1.3 1.2 0.7 1.33.3 0.6 1.6 Rifabutin Inhibition of RNA- Antibacterial polymeraseDNA-dependent 1.1 1.0 1.5 1.3 1.6 1.1 1.4 Torsemide Diuretic 0.7 0.6 0.70.7 0.8 0.7 1.2 Parbendazole Anthelmintic (VET) 1.0 1.0 1.2 1.1 1.3 1.11.3 Halofantrine hydrochloride Antimalarial 1.4 1.1 1.4 1.0 1.2 1.1 1.6Mecamylamine hydrochloride Antihypertensor 1.1 1.2 1.1 1.1 1.2 1.0 1.6Articaine hydrochloride Local Anesthesic 1.0 1.0 1.1 1.1 1.1 0.9 1.7Procarbazine hydrochloride DNA depolymerization 1.1 1.2 1.0 1.1 1.1 1.11.5 Nomegestrol acetate Progestin 1.0 1.0 1.3 0.9 1.1 0.9 1.6 Viomycinsulfate Antibacterial 1.1 1.0 1.1 1.0 1.1 0.9 1.6 Pancuronium bromideCompetitive antagonist of Neuromuscular blocking agent autonomiccholinergic receptors 1.3 1.2 0.9 1.1 1.5 1.0 1.8 Saquinavir mesylateProtease inhibitor 1.1 1.0 1.1 1.0 1.1 0.9 1.7 Molindone hydrochlorideAntipsychotic 1.1 1.1 1.3 1.7 1.6 1.0 1.6 Ronidazole Antimicrobial 1.41.0 1.2 1.0 1.4 1.0 2.2 Alcuronium chloride Neuromuscular blocking agent1.1 2.3 1.0 1.1 1.2 0.9 1.3 Dorzolamide hydrochloride Antiglaucoma drug1.3 1.1 1.1 1.1 1.2 1.0 1.8 Zalcitabine Reverse transcriptase Antiviral(HIV) inhibitor 1.1 1.0 1.2 1.0 1.2 1.0 1.9 Azaperone Tranquilizer 1.21.1 1.2 1.2 1.4 1.1 1.8 Methyldopate hydrochloride Antihypertensor 19.31.0 3.1 4.9 3.4 15.6 1.9 Cefepime hydrochloride Antibacterial 1.1 1.01.1 1.1 1.2 1.0 1.6 Levocabastine hydrochloride Antihistaminic 1.0 1.11.1 1.0 1.2 1.0 1.6 Clocortolone pivalate Glucocorticoid 0.8 1.5 1.4 0.80.9 0.7 13.2 Pyrvinium pamoate Anthelmintic (Nematodes, enterobiasis)13.4 2.8 4.3 5.2 2.9 17.7 1.7 Nadifloxacin Antibacterial

Consistent with expectations, a detailed assessment of the hits revealedthat AK screening enriches for the identification of bactericidalcompounds (Table 3). More specifically, 100% of the antibiotics thatwere identified to be active against P. aeruginosa or K. pneumoniaerepresented bactericidal agents. Similarly, 96%, 94%, and 80% of theantibiotics that were active against E. coli, E. cloacae, and E.faecium, respectively, were bactericidal antibiotics. The assayidentified 71% and 64% bactericidal antibiotics for A. baumannii and S.aureus.

Among the bactericidal antibiotics detected, β-lactam, cephalosporin,polymyxin, and fluoroquinolone antibiotics were active againstGram-negative pathogens (E. coli and the ESKAPE pathogens A. baumannii,P. aeruginosa, and K. pneumoniae), and penicillins, cephalosporins,quinolines, glycopeptides, and carbapenems were active toward S. aureus(see Table 4). It was also determined that clofazimine, which wasinitially developed as an antimycobacterial agent and was recently shownto exhibit bactericidal activity toward S. aureus, was indeed activeagainst S. aureus. Bacteriostatic compounds, such as clindamycin, andmacrolides, such as erythromycin, were not identified as being activetoward any of the species tested. Interestingly, many tetracyclines wereidentified as killing S. aureus strain USA300-0114 and A. baumanniistrain 98-37-09. In addition, the library contains membrane-activeantiseptics, such as chlorhexidine, and with the exception of E.cloacae, these were also strong hits against all organisms tested.Screening results also revealed that the E. faecium strain tested provedto be resistant to most classes of antibiotics within the Prestwicklibrary by MIC testing (Table 2), and this was also observed in the AKassay, indicating that the assay exhibits a low false-positive rate.Interestingly, the assay also detected 38 drugs with no knownantimicrobial properties (denoted as “other” in Table 3) that wereactive against each organism; see Table 4 for a complete list of thesecompounds.

Antimicrobial Activities of Nonantibiotic Drugs that Induce AK Release—

Recently, the exploration of the so-called “off-target” activities ofpreviously developed drugs has emerged as an approach to identifychemical scaffolds that could be exploited for new therapeuticindications. In that regard, Prestwick library screening resultsrevealed that 4% to 56% (depending on the organism) of the members thatgenerated significant AK signal were compounds with no previouslyreported antimicrobial activity (see Table 4).

To determine whether the 38 nonantibiotics identified in the screen havepotential for repurposing as anti-infectives, 4 nonantibiotic drugs thatwere commercially available were further evaluated by two secondaryassays. First, dose-response assays were performed to validate that theyinduced AK activity, and all were reconfirmed. Second, the in vitroantimicrobial activity for each drug was measured by standard MICtesting. With the exception of one drug/organism pair, all drugsexhibited in vitro activity toward each organism. More specifically,tamoxifen, suloctidil, and clomiphene exhibited MICs of 8 μg ml⁻¹against E. faecium. S. aureus and A. baumannii were susceptible toterfenadine (16-μg ml⁻¹ MIC and 64-μg ml⁻¹ MIC, respectively).Suloctidil was also detected to be active against P. aeruginosa by bothprimary and confirmatory AK screens, but the drug did not elicit anantimicrobial response by MIC measures. Some strains of P. aeruginosasecrete AK at high cell density, and it is possible that suloctidil maytrigger a similar response.

Terfenadine and tamoxifen, which exhibited antimicrobial propertiestoward planktonic S. aureus and E. faecium cells, respectively, werecharacterized further. Terfenadine was evaluated for activity against S.aureus biofilms and small-colony variants using the AK assays describedabove. Treatment of 48-h S. aureus strain UAMS-1 biofilms with 10×-MICterfenadine elicited a modest 2.7-fold increase in AK release (see FIG.6A); plating-based viability assays determined that this correlated witha 1.1-log reduction in biofilm cell viability, which was comparable tothe activity of ciprofloxacin under the same assay conditions (see FIG.4A). Similarly, treatment of S. aureus small-colony-variant UAMS-1112cells with 10×MIC terfenadine elicited a 3.3-fold increase in AK signalin comparison to that for mock-treated cells (see FIG. 6A). ConventionalMIC testing subsequently verified that terfenadine is active againstUAMS-1112 at 2 μg ml⁻¹.

Next, the in vivo antimicrobial properties of terfenadine and tamoxifenwere evaluated using a Galleria mellonella model of S. aureus and E.faecium infection, respectively. For terfenadine, groups of larvae(n=45) were infected with 1.0×10⁶ S. aureus USA300-1114 cells. Wormswere then treated at 2 h and 24 h with a range of terfenadineconcentrations (20 to 160 mg kg⁻¹), vehicle (DMSO; negative control), or20 mg kg⁻¹ vancomycin (positive control), and larval survival wasassessed 48 h postinoculation. For tamoxifen studies, experiments wereperformed exactly as described above except that larvae were inoculatedwith 1.4×10⁶ E. faecium strain 824-05 cells and larvae were treated witheither 80 or 160 mg kg⁻¹ kg tamoxifen. Terfenadine-treated larvae didnot reproducibly exhibit increased survival relative to vehicle-treatedlarvae. However, tamoxifen treatment of E. faecium-infected larvaeresulted in a dose-dependent increase in survival. As shown in FIG. 6B,treatment with 80 or 160 mg kg⁻¹ resulted in a 4.2-fold- or7-fold-higher survival, respectively, than was found withvehicle-treated controls. Taken together, these results indicate thatterfenadine exhibits in vitro activity toward S. aureus planktonic, SCV,and biofilm populations and that tamoxifen is active toward planktonicE. faecium and against the organism in a simple animal model ofinfection. As discussed below, these data show that tamoxifen andterfenadine represent attractive new chemical scaffolds forantibacterial optimization.

As set forth herein, Prestwick library screening revealed that eachbacterial species studied was susceptible to members of the library withno previously reported antimicrobial activity. The antimicrobialproperties of two of these drugs was studied in more detail: terfenadineand tamoxifen. Terfenadine is a nonsedating antihistamine based on a4-substituted piperidine scaffold. Based on further studies, it wasdetermined that terfenadine acts as a topoisomerase inhibitor and isstructurally similar to certain topoisomerase inhibitors, including forexample novel bacterial topoisomerase II inhibitor (NBTI). Based on thestructural similarity between terfenadine and these molecules, it islikely that terfenadine is acting as a DNA gyrase and topoisomeraseinhibitor. More specifically, terfenadine and derivatives thereof act astopoisomerase 4 inhibitors. Furthermore, it was found that terfenadinehas activity against S. aureus small-colony variants and biofilms,properties not previously reported for this scaffold.

A screen of E. faecium identified two structurally related nonsteroidestrogen receptor antagonists, tamoxifen and clomiphene. Tamoxifen isused to treat some forms of estrogen-receptor-positive breast cancer,while clomiphene is used in fertility treatment regimens. These twocompounds are members of the triarylethylene class of estrogenreceptors.

Since the number of agents with activity toward enterococcus is quitelimited, tamoxifen's in vivo activity was investigated using a Galleriamodel of enterococcus infection. Although it was not as active asvancomycin, tamoxifen did impart a survival benefit, indicating that ithas in vivo antimicrobial activity. High-dose tamoxifen therapy has beenused in experimental treatment of refractory human cancers, and dosingresults in micromolar serum concentrations of tamoxifen corresponding tothe levels of drug associated with antienterococcal activity observed inthe studies provided herein.

In addition to providing a powerful new HTS approach to identifyantimicrobial agents active against planktonic bacteria, an AK assaythat is easily amenable to screening bacteria in disease states thatcannot be readily screened via conventional approaches is providedherein. In that regard, the AK assay is capable of measuring the killingproperties of bactericidal agents administered to biofilms formed byboth Gram-negative and Gram-positive representatives of the ESKAPEpathogens. Similarly, the AK assay can detect the bactericidalproperties of antibiotics toward a phenotypically stable S. aureussmall-colony variant. These features can be exploited to developcorresponding high-throughput screening assays for the identification ofagents that kill established biofilms and small-colony variants orprovide powerful secondary assays aimed at characterizing the potentialantimicrobial properties of molecules of interest.

Example II Antimicrobial Properties of Terfenadine and TerfenadineDerivatives

Terfenadine derivatives were synthesized as described below.

General Method A KSC-335-0071-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one

To a vial was added the diphenyl(piperidin-4-yl)methanol (1.190 g, 4.45mmol), 1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-one (1.012 g, 4.24mmol), sodium bicarbonate (0.427 g, 5.09 mmol) with water:2-butanone (18mL, 1:5). The reaction stirred at 85° C. for 16 h and was then cooled tort and water (50 mL) was added. The reaction was extracted with EtOAc(3×50 mL). The EtOAc layer was dried with MgSO₄, filtered, concentratedand purified by MPLC (20 min, 0-10% MeOH:DCM) to produce pure1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one (1.37 g, 2.92 mmol, 69% yield) as a colorlessoil. ¹H NMR (500 MHz, CDCl₃): δ 7.90 (d, J=8.4 Hz, 2H), 7.49-7.45 (m,6H), 7.31-7.25 (m, 4H), 7.20-7.14 (m, 2H), 2.97-2.92 (m, 4H), 2.45-2.36(m, 3H), 2.09 (br s, 1H), 2.00-1.87 (m, 4H), 1.49-1.32 (m, 4H), 1.34 (s,9H). ¹³C NMR (125 MHz, CDCl₃): δ 199.7, 156.5, 146.0, 134.5, 128.1,128.0, 126.4, 125.7, 125.4, 79.4, 57.9, 43.9, 43.4, 44.1, 36.2, 35.0,31.0, 26.2, 21.9. LCMS Retention time: 4.207 min. LCMS purity 99.5%.HRMS (ESI): m/z calcd for C₃₂H₃₉NO₂ [M+H]⁺ 470.2981. found 470.3054.

KSC-335-0054-(4-(Hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-isopropylphenyl)butan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.620 g, 2.317 mmol),4-chloro-1-(4-isopropylphenyl)butan-1-one (0.496 g, 2.207 mmol), sodiumbicarbonate (0.222 g, 2.65

mmol) with water (3 mL) and 2-butanone (15 mL, Ratio: 5). to producepure4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-isopropylphenyl)butan-1-one(0.481 g, 1.056 mmol, 48% yield) as a colorless oil. ¹H NMR (500 MHz,CDCl₃): δ 7.89 (d, J=8.4 Hz, 2H), 7.49-7.45 (m, 4H), 7.31-7.26 (m, 6H),7.20-7.14 (m, 2H), 2.98-2.91 (m, 4H), 2.45-2.35 (m, 1H), 2.38 (t, J=6.8Hz, 2H), 2.08 (br s, 1H), 1.99-1.89 (m, 4H), 1.61 (br s, 1H), 1.48-1.33(m, 4H), 1.27 (d, J=6.8 Hz, 6H).

KSC-335-0061-(4-(tert-butyl)phenyl)-5-(4-(hydroxydiphenylmethyl)piperidin-1-yl)pentan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.544 g, 2.035 mmol),1-(4-(tert-butyl)phenyl)-5-chloropentan-1-one (0.490 g, 1.938 mmol),sodium bicarbonate (0.195 g, 2.326 mmol) with water:2-butanone (18 mL,1:5) to produce pure1-(4-(tert-butyl)phenyl)-5-(4-(hydroxydiphenylmethyl)piperidin-1-yl)pentan-1-one(0.600 g, 1.240 mmol, 64.0% yield) as a colorless oil. ¹H NMR (500 MHz,CDCl₃): δ 7.89 (d, J=8.4 Hz, 2H), 7.49-7.45 (m, 6H), 7.32-7.26 (m, 4H),7.20-7.14 (m, 2H), 2.98-2.92 (m, 4H), 2.48-2.32 (m, 3H), 2.14 (br s,1H), 1.99-1.89 (m, 2H), 1.77-1.62 (m, 2H), 1.60-1.42 (m, 6H), 1.34 (s,9H).

KSC-335-0081-(4-(tert-butyl)phenyl)-3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.543 g, 2.032 mmol),1-(4-(tert-butyl)phenyl)-3-chloropropan-1-one (0.435 g, 1.936 mmol),sodium bicarbonate (0.195 g, 2.323 mmol) with water:2-butanone (18 mL,1:5) to produce pure 1-(4-(tert-butyl)phenyl)-3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propan-1-one (0.838 g, 1.84 mmol, 95% yield) as acolorless oil. ¹H NMR (500 MHz, CDCl₃): δ 7.89 (d, J=8.4 Hz, 2H),7.50-7.45 (m, 6H), 7.32-7.25 (m, 4H), 7.21-7.14 (m, 2H), 3.15 (t, J=7.1Hz, 2H), 3.01-2.96 (m, 2H), 2.81 (t, J=7.1 Hz, 2H), 2.49-2.42 (m, 1H),2.15-2.04 (m, 3H), 1.56-1.46 (m, 4H), 1.33 (s, 9H). ¹³C NMR (125 MHz,CDCl₃): δ 198.9, 171.1, 156.8, 145.8, 134.3, 128.2, 128.0, 126.5, 125.8,125.5, 79.4, 60.4, 54.2, 53.4, 44.0, 36.3, 35.1, 31.1, 26.4, 21.1, 14.2.LCMS Retention time: 3.968 min. LCMS purity 98.1%. HRMS (ESI): m/z calcdfor C₃₁H₃₇NO₂ [M+H]⁺ 456.2824. found 456.2897.

KSC-335-0091-(4-chlorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.626 g, 2.341 mmol),4-chloro-1-(4-chlorophenyl) butan-1-one (0.484 g, 2.229 mmol), sodiumbicarbonate (0.225 g, 2.68 mmol) with water:2-butanone (18 mL, 1:5) toproduce pure1-(4-chlorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(0.394 g, 0.879 mmol, 39.4% yield) as a colorless oil. ¹H NMR (400 MHz,CDCl₃): δ 7.90 (d, J=8.4 Hz, 2H), 7.48-7.44 (m, 4H), 7.42 (d, J=8.6 Hz,2H), 7.31-7.26 (m, 4H), 7.20-7.15 (m, 2H), 3.96-2.89 (m, 4H), 2.45-2.34(m, 3H), 2.08 (br s, 1H), 1.99-1.87 (m, 4H), 1.50-1.30 (m, 4H).

General Method B KSC-335-014, Terfenadine1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol

To a vial was added the1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(KSC-335-007 (0.198 g, 0.422 mmol) and MeOH (2 mL). The sodiumborohydride (0.032 g, 0.844 mmol) was then added and the reactionstirred at rt for 3 h. The reaction was concentrated to dryness, water(5 mL) was added and a white precipitate formed. The precipitate wasfiltered out and then dissolved in DCM (10 mL), dried with MgSO₄,filtered and concentrated to produce pure1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol(0.151 g, 0.320 mmol, 76% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.52-7.46 (m, 4H), 7.33-7.25 (m, 8H), 7.21-7.15 (m, 2H), 4.61-4.56 (m,1H), 3.16-3.11 (br m, 1H), 3.00-2.94 (m, 1H), 2.51-2.34 (m, 4H),2.10-1.88 (m, 4H), 1.83-1.75 (m, 1H), 1.70-1.45 (m, 6H), 1.30 (s, 9H).¹³C NMR (125 MHz, CDCl₃): δ 149.4, 146.1, 146.0, 142.7, 128.2, 128.1,126.4, 126.3, 125.7, 125.6, 125.3, 125.0, 79.2, 73.4, 58.9, 54.7, 53.3,44.2, 39.7, 34.4, 31.4, 26.0, 25.9, 24.1. LCMS Retention time: 4.137min. LCMS purity 97.5%. HRMS (ESI): m/z calcd for C₃₂H₄₁NO₂ [M+H]⁺472.3144. found 472.3219.

KSC-335-0124-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-isopropylphenyl)butan-1-ol

Method B:4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-isopropylphenyl)butan-1-one(KSC-335-005) (0.154 g, 0.338 mmol) and MeOH (2 mL) sodium borohydride(0.026 g, 0.676 mmol) to produce pure4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-isopropylphenyl)butan-1-ol(0.146 g, 0.319 mmol, 94% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.52-7.46 (m, 4H), 7.32-7.23 (m, 6H), 7.20-7.14 (m, 4H), 4.61-4.58 (m,1H), 3.18-3.11 (br m, 1H), 3.00-2.94 (br m, 1H), 2.87 (septet, J=6.9 Hz,1H), 2.50-2.33 (m, 3H), 2.29 (br s, 1H), 2.10-1.89 (m, 3H), 1.82-1.44(m, 8H), 1.22 (d, J=6.9 Hz, 6H). ¹³C NMR (125 MHz, CDCl₃): δ 147.2,146.1, 146.0, 128.2, 128.1, 126.5, 126.4, 126.1, 125.7, 125.7, 79.2,73.5, 58.9, 54.7, 53.3, 44.2, 39.9, 33.7, 26.0, 25.9, 24.2, 24.1, 24.0.LCMS Retention time: 4.056 min. LCMS purity 98.6%. HRMS (ESI): m/z calcdfor C₃₁H₃₉NO₂ [M+H]⁺ 458.2986. found 458.3062.

KSC-335-0131-(4-(tert-butyl)phenyl)-5-(4-(hydroxydiphenylmethyl)piperidin-1-yl)pentan-1-ol

Method B:1-(4-(tert-butyl)phenyl)-5-(4-(hydroxydiphenylmethyl)piperidin-1-yl)pentan-1-one(KSC-335-006) (0.204 g, 0.422 mmol) and MeOH (2 mL) and sodiumborohydride (0.032 g, 0.844 mmol) to produce pure1-(4-(tert-butyl)phenyl)-5-(4-(hydroxydiphenylmethyl)piperidin-1-yl)pentan-1-ol(0.156 g, 0.321 mmol, 76% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.50-7.45 (m, 4H), 7.36 (d, J=8.4 Hz, 2H), 7.31-7.24 (m, 6H), 7.19-7.14(m, 2H), 4.65-4.60 (m, 1H), 2.98-2.90 (br m, 1H), 2.48-2.38 (m, 1H),2.30 (t, J=7.2 Hz, 2H), 2.23 (br s, 1H), 1.97-1.87 (m, 2H), 1.84-1.60(m, 4H), 1.55-1.35 (m, 8H), 1.31 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ150.3, 146.0, 142.0, 128.1, 126.4, 125.8, 125.5, 125.3, 79.4, 74.1,58.4, 54.1, 54.0, 44.2, 38.5, 34.5, 31.4, 31.3, 26.5, 26.3, 26.2, 23.7.LCMS Retention time: 4.254 min. LCMS purity 96.4%. HRMS (ESI): m/z calcdfor C₃₃H₄₃NO₂ [M+H]⁺ 486.3294. found 486.3370.

KSC-335-0151-(4-chlorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol

Method B:1-(4-chlorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(KSC-335-009) (0.156 g, 0.348 mmol) and MeOH (2 mL) sodium borohydride(0.026 g, 0.696 mmol) to produce pure1-(4-chlorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol(0.118 g, 0.262 mmol, 75% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.51-7.46 (m, 4H), 7.32-7.24 (m, 8H), 7.20-7.15 (m, 2H), 4.61-4.56 (m,1H), 3.16-3.10 (br m, 1H), 2.97-2.91 (m, 1H), 2.51-2.33 (m, 4H),2.12-1.88 (m, 3H), 1.83-1.75 (m, 1H), 1.78-1.46 (m, 8H). ¹³C NMR (125MHz, CDCl₃): δ 146.0, 145.9, 144.5, 139.1, 128.19, 128.18, 128.1, 127.1,126.49, 126.45, 125.62, 125.57, 79.2, 72.9, 58.8, 54.7, 53.2, 44.2,40.2, 26.0, 25.9, 24.1. LCMS Retention time: 3.845 min. LCMS purity98.4%. HRMS (ESI): m/z calcd for C₂₈H₃₂ClNO₂ [M+H]⁺ 450.2122. found450.2194.

KSC-335-0161-(4-(tert-butyl)phenyl)-3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propan-1-ol

Method B:1-(4-(tert-butyl)phenyl)-3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propan-1-one(KSC-335-008) (0.116 g, 0.255 mmol) and MeOH (2 mL) and sodiumborohydride (0.019 g, 0.509 mmol) to produce pure1-(4-(tert-butyl)phenyl)-3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propan-1-ol(0.106 g, 0.232 mmol, 91% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.49-7.44 (m, 4H), 7.36-7.25 (m, 8H), 7.22-7.16 (m, 2H), 6.72 (br s,1H), 4.90-4.85 (m, 1H), 3.21-3.15 (br m, 1H), 3.11-3.05 (br m, 1H),2.70-2.62 (m, 1H), 2.57-2.40 (m, 2H), 2.14-2.06 (m, 2H), 1.91-1.79 (m,3H), 1.57-1.45 (m, 4H), 1.31 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ 149.6,145.8, 145.7, 141.9, 128.2, 128.1, 126.6, 126.5, 125.8, 125.7, 125.2,125.0, 79.4, 75.3, 57.3, 55.2, 53.2, 44.1, 34.4, 33.7, 31.4, 26.7, 26.4.LCMS Retention time: 4.006 min. LCMS purity 97.7%. HRMS (ESI): m/z calcdfor C₃₁H₃₉NO₂ [M+H]⁺ 458.2991. found 458.3066.

KSC-335-018

4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-phenylbutan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.524 g, 1.961 mmol),4-chloro-1-phenylbutan-1-one (0.300 ml, 1.868 mmol), sodium bicarbonate(0.188 g, 2.241 mmol) with water:2-butanone (18 mL, 1:5) to produce pure4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-phenylbutan-1-one (0.177g, 0.428 mmol, 23% yield) as a colorless oil. ¹H NMR (500 MHz, CDCl₃): δ7.98-7.94 (m, 2H), 7.57-7.52 (m, 1H), 7.49-7.42 (m, 6H), 7.32-7.26 (m,4H), 7.20-7.15 (m, 2H), 3.00-2.90 (m, 4H), 2.46-2.35 (m, 3H), 2.09 (brs, 1H), 2.00-1.87 (m, 4H), 1.50-1.33 (m, 4H).

KSC-335-020 1-(tert-butyl)-4-(4-chlorobutyl)benzene

To a vial was added the 1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-one(0.266 g, 1.114 mmol) and triethylsilane (0.518 g, 0.712 ml, 4.46 mmol)with TFA (4 mL). The reaction stirred at 75° C. for 18 h, was cooled tort and concentrated in vacuo. The residue was then dissolved in DCM (5mL) and washed with water (4 mL). The DCM layer was collected and washedwith water (1×5 mL), dried with MgSO₄, filtered and adsorbed to silicaand purified by Teledyne ISCO Combiflash chromatography (20 min, 0-40%EtOAc:Hex) and fractions 4 and 5 were collected to produce pure1-(tert-butyl)-4-(4-chlorobutyl)benzene (0.155 g, 0.690 mmol, 61.9%yield) as an oil. ¹H NMR (400 MHz, CDCl₃): δ 7.32 (d, J=7.6 Hz, 2H),7.12 (d, J=7.6 Hz, 2H), 3.56 (t, J=6.5 Hz, 2H), 2.62 (t, J=7.5 Hz, 2H),1.87-1.74 (m, 4H), 1.32 (s, 9H).

KSC-335-0214-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-phenylbutan-1-ol

Method B:4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-phenylbutan-1-one(KSC-335-018) (0.065 g, 0.157 mmol) and MeOH (2 mL) and sodiumborohydride (0.012 g, 0.314 mmol) to produce pure4-(4-(hydroxydiphenylmethyl) piperidin-1-yl)-1-phenylbutan-1-ol (0.048g, 0.116 mmol, 73% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.51-7.46 (m, 4H), 7.37-7.33 (m, 2H), 7.32-7.26 (m, 5H), 7.23-7.14 (m,3H), 4.65-4.61 (m, 1H), 3.15 (d, J=11.7 Hz, 1H), 2.96 (d, J=11.7 Hz,1H), 2.52-2.31 (m, 4H), 2.11-1.90 (m, 3H), 1.84-1.74 (m, 2H), 1.70-1.44(m, 7H). ¹³C NMR (125 MHz, CDCl₃): δ 146.0, 145.9, 145.8, 128.2, 128.1,128.0, 126.6, 126.5, 126.4, 125.6, 125.6, 79.2, 73.6, 58.9, 54.7, 53.3,44.2, 40.1, 26.0, 25.9, 24.1. LCMS Retention time: 3.711 min. LCMSpurity 99.8%. HRMS (ESI): m/z calcd for C₂₈H₃₃NO₂ [M+H]⁺ 416.2517. found416.2592.

KSC-335-022 Methyl2-methyl-2-(4-(4-((methylsulfonyl)oxy)but-1-yn-1-yl)phenyl)propanoate

To a vial was added the methyl2-(4-(4-hydroxybut-1-yn-1-yl)phenyl)-2-methylpropanoate (0.051 g, 0.207mmol) and dry DCM (2 mL). The methanesulfonyl chloride (0.047 g, 0.032mL, 0.414 mmol) and pyridine (0.147 g, 0.151 mL, 1.864 mmol) were eachadded and the reaction and stirred at rt for 16 h. The reaction was thendiluted with DCM (5 mL) and washed with 1% w/v sulfuric acid in water(3×7 mL), saturated NaHCO₃ (7 mL) and brine (7 mL). The organic layerwas dried with MgSO₄, filtered and concentrated to produce methyl2-methyl-2-(4-(4-((methylsulfonyl) oxy)but-1-yn-1-yl)phenyl)propanoate(0.063 g, 0.194 mmol, 94% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.35 (d,J=8.6 Hz, 2H), 7.26 (d, J=8.6 Hz, 2H), 4.38 (t, J=6.8 Hz, 2H), 3.64 (s,3H), 3.06 (s, 3H), 2.87 (t, J=6.8 Hz, 2H), 1.56 (s, 6H).

KSC-335-0234-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-methoxyphenyl)butan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.490 g, 1.832 mmol),4-chloro-1-(4-methoxyphenyl)butan-1-one (0.371 g, 1.744 mmol), sodiumbicarbonate (0.176 g, 2.093 mmol) with water (3 mL) and 2-butanone (15mL) to produce pure4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-methoxyphenyl)butan-1-one(0.282 g, 0.636 mmol, 36% yield) as a colorless oil. ¹H NMR (400 MHz,CDCl₃): δ 7.94 (d, J=9.0 Hz, 2H), 7.49-7.45 (m, 4H), 7.31-7.26 (m, 4H),7.17 (tt, J₁=7.3 Hz, J₂=1.3 Hz, 2H), 6.92 (d, J=8.9 Hz, 2H), 3.87 (s,3H), 2.97 (br s, 1H), 2.93 (t, J=7.3 Hz, 2H), 2.48-2.35 (m, 3H),2.02-1.88 (m, 4H), 1.63-1.40 (m, 4H).

KSC-335-0241-(4-Fluorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.385 g, 1.439 mmol),4-chloro-1-(4-fluorophenyl)butan-1-one (0.275 g, 1.371 mmol), sodiumbicarbonate (0.138 g, 1.645 mmol) with water (3 mL) and 2-butanone (15mL) to produce pure1-(4-fluorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(0.174 g, 0.403 mmol, 29% yield) as a colorless oil. ¹H NMR (400 MHz,CDCl₃): δ 8.00-7.96 (m, 2H), 7.48-7.44 (m, 4H), 7.31-7.26 (m, 4H),7.20-7.15 (m, 2H), 7.14-7.08 (m, 2H), 2.97-2.94 (m, 4H), 2.46-2.36 (m,3H), 2.15 (br s, 1H), 2.01-1.88 (m, 4H), 1.52-1.35 (m, 4H).

KSC-335-0251-(4-Bromophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one

Method A: diphenyl(piperidin-4-yl)methanol (0.439 g, 1.642 mmol),1-(4-bromophenyl)-4-chlorobutan-1-one (0.409 g, 1.564 mmol), sodiumbicarbonate (0.158 g, 1.877 mmol) with water (3 mL) and 2-butanone (15mL) to produce pure1-(4-bromophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(0.245 g, 0.498 mmol, 32% yield) as a colorless oil. ¹H NMR (400 MHz,CDCl₃): δ 7.83-7.79 (m, 2H), 7.60-7.56 (m, 2H), 7.48-7.45 (m, 4H),7.31-7.26 (m, 4H), 7.20-7.15 (m, 2H), 2.95-2.90 (m, 4H), 2.45-2.35 (m,3H), 2.17 (br s, 1H), 2.00-1.88 (m, 4H), 1.51-1.33 (m, 4H).

KSC-335-0304-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-methoxyphenyl)butan-1-ol

Method B:4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-methoxyphenyl)butan-1-one(KSC-335-023) (0.113 g, 0.255 mmol) and MeOH (2 mL) and sodiumborohydride (0.019 g, 0.509 mmol) to produce pure4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(4-methoxyphenyl)butan-1-ol(0.062 g, 0.139 mmol, 55% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.51-7.46 (m, 4H), 7.32-7.24 (m, 6H), 7.20-7.15 (m, 2H), 6.84 (d, J=7.0Hz, 2H), 4.60-4.56 (m, 1H), 3.78 (s, 3H), 3.14 (d, J=11.7 Hz, 1H), 2.95(d, J=11.7 Hz, 1H), 2.50-2.35 (m, 4H), 2.10-2.02 (m, 1H), 1.99-1.86 (m,2H), 1.80-1.73 (m, 1H), 1.69-1.45 (m, 7H). ¹³C NMR (125 MHz, CDCl₃): δ158.3, 146.1, 146.0, 138.1, 128.1, 128.1, 126.7, 126.4, 126.4, 125.7,125.6, 113.5, 79.2, 73.2, 58.9, 55.2, 54.7, 53.2, 44.2, 40.0, 26.0,25.9, 24.1. LCMS Retention time: 3.620 min. LCMS purity 98.3%. HRMS(ESI): m/z calcd for C₂₉H₃₅NO₃ [M+H]⁺ 446.2652. found 446.2728.

KSC-335-0311-(4-fluorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol

Method B:1-(4-fluorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(KSC-335-024) (0.064 g, 0.148 mmol) and MeOH (2 mL) and sodiumborohydride (0.011 g, 0.297 mmol) to produce pure1-(4-fluorophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol(0.059 g, 0.136 mmol, 92% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.51-7.46 (m, 4H), 7.33-7.25 (m, 6H), 7.20-7.14 (m, 2H), 6.96 (t, J=8.8Hz, 2H), 4.61-4.57 (m, 1H), 3.14 (d, J=11.7 Hz, 1H), 2.94 (d, J=11.7 Hz,1H), 2.51-2.35 (m, 4H), 2.13-2.02 (m, 1H), 2.02-1.86 (m, 2H), 1.78-1.45(m, 7H). ¹³C NMR (125 MHz, CDCl₃): δ 162.6, 160.7, 146.0, 145.9, 141.7,141.6, 128.2, 128.1, 127.2, 127.1, 126.5, 126.4, 125.6, 125.5, 114.9,114.7, 79.2, 73.0, 58.8, 54.8, 53.1, 44.2, 40.3, 30.9, 26.0, 25.9, 24.1.LCMS Retention time: 3.691 min. LCMS purity 98.5%. HRMS (ESI): m/z calcdfor C₂₈H₃₂FNO₂ [M+H]⁺ 434.2406. found 434.2482.

KSC-335-0321-(4-bromophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol

Method B:1-(4-bromophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(KSC-335-025) (0.083 g, 0.169 mmol) and MeOH (2 mL) and sodiumborohydride (0.013 g, 0.337 mmol) to produce pure1-(4-bromophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol(0.053 g, 0.107 mmol, 63.6% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.51-7.46 (m, 4H), 7.41 (d, J=6.8 Hz, 2H), 7.32-7.26 (m, 4H), 7.22 (d,J=6.6 Hz, 2H), 7.20-7.15 (m, 2H), 4.60-4.56 (m, 1H), 3.14 (d, J=11.7 Hz,1H), 2.93 (d, J=11.7 Hz, 1H), 2.50-2.42 (m, 1H), 2.41-2.34 (m, 2H),2.11-2.05 (m, 1H), 2.01-1.95 (m, 1H), 1.94-1.86 (m, 1H), 1.78-1.45 (m,8H). ¹³C NMR (125 MHz, CDCl₃): δ 146.0, 145.9, 145.1, 131.1, 128.2,128.1, 127.5, 126.5, 126.4, 125.6, 125.6, 120.2, 79.2, 72.9, 58.8, 54.7,53.2, 44.1, 40.1, 26.0, 25.9, 24.1. LCMS Retention time: 3.902 min. LCMSpurity 99.1%. HRMS (ESI): m/z calcd for C₂₈H₃₂BrNO₂ [M+H]⁺ 494.1614.found 494.1673.

KSC-335-041(1-(4-(4-(tert-butyl)phenyl)butyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.153 g, 0.571mmol), 1-(tert-butyl)-4-(4-chlorobutyl)benzene (KSC-335-020) (0.154 g,0.685 mmol) and potassium carbonate (0.473 g, 3.43 mmol) inacetonitrile. The reaction stirred overnight at 85° C. and for 18 h andwas then cooled to rt and filtered. The filtrate was then diluted withbrine and extracted with diethyl ether (3×15 mL). The ether layers werecombined, dried with MgSO₄, filtered and purified by reverse-phase MPLC(20 min, 10-100% MeCN:H₂O) to produce pure(1-(4-(4-(tert-butyl)phenyl)butyl)piperidin-4-yl)diphenylmethanol (0.180g, 0.395 mmol, 69% yield) as an oil. ¹H NMR (400 MHz, CDCl₃): δ7.38-7.34 (m, 4H), 7.19-7.14 (m, 6H), 7.07-7.02 (m, 2H), 6.99-6.95 (m,2H), 2.86-2.80 (m, 2H), 2.45 (t, J=7.3 Hz, 2H), 2.35-2.26 (m, 1H),2.22-2.17 (m, 2H), 1.84-1.76 (m, 3H), 1.52-1.30 (m, 8H), 1.18 (s, 9H).¹³C NMR (125 MHz, CDCl₃): δ 171.1, 148.4, 146.0, 139.4, 128.1, 128.0,126.4, 125.8, 125.1, 79.5, 60.4, 58.8, 54.1, 44.2, 35.2, 34.3, 31.4,29.5, 26.8, 26.4, 21.0, 14.2. LCMS Retention time: 3.928 min. LCMSpurity 97.8%. HRMS (ESI): m/z calcd for C₃₂H₄₁NO [M+H]⁺ 456.3188. found456.3261.

KSC-335-053 Methyl 4-(1,3-dithian-2-yl)benzoate

A flame dried vial was evaporated 3 times with argon and methyl4-formylbenzoate (0.50 g, 3.05 mmol) was added with anhydrous DCM (8.70mL) followed by 1,3-propanedithiol (0.339 mL, 3.35 mmol). The reactionbegan to stir at rt for 1.5 h. The reaction was then cooled to 0° C. andthe BF3.OEt₂ (0.425 ml, 3.35 mmol) was added dropwise. The reaction wasthen warmed slowly to rt and stirred overnight. The reaction was thendiluted with DCM (15 mL) and quenched with saturated NaHCO₃ (15 mL) andthe DCM layer was dried with MgSO₄, filtered and adsorbed to silica andpurified by MPLC (20 min, 0-35% EtOAc:Hex) to produce methyl4-(1,3-dithian-2-yl)benzoate (0.669 g, 2.63 mmol, 86% yield). ¹H NMR(400 MHz, CDCl₃): δ 8.01 (d, J=8.4 Hz, 2H), 7.54 (d, J=8.3 Hz, 2H), 3.91(s, 3H), 3.12-3.03 (m, 2H), 2.96-2.90 (m, 2H), 2.23-2.16 (m, 1H),2.01-1.89 (m, 1H), 1.55 (s, 2H).

KSC-335-054 Methyl2-(4-(4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)but-1-yn-1-yl)phenyl)-2-methylpropanoate

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.528 g, 1.974mmol), methyl2-methyl-2-(4-(4-((methylsulfonyl)oxy)but-1-yn-1-yl)phenyl)propanoate(KSC-335-022) (0.582 g, 1.794 mmol) and potassium carbonate (0.744 g,5.38 mmol) with acetonitrile (10 mL). The reaction stirred at 70° C. for18 h and cooled to rt and filtered to remove the potassium carbonate.The filtrate was adsorbed to silica gel and purified by reverse-phaseMPLC (20 min, 10-100% MeCN:H₂O) to produce pure methyl2-(4-(4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)but-1-yn-1-yl)phenyl)-2-methylpropanoate (0.434 g, 0.876mmol, 49% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.49-7.46 (m, 4H),7.34-7.27 (m, 6H), 7.25-7.15 (m, 4H), 3.64 (s, 3H), 3.03-2.97 (m, 2H),2.68-2.63 (m, 2H), 2.59-2.54 (m, 2H), 2.48-2.40 (m, 1H), 2.14-2.06 (m,2H), 1.63 (br s, 1H), 1.55 (s, 6H), 1.55-1.45 (m, 4H).

KSC-335-056 Methyl 4-(2-(3-chloropropyl)-1,3-dithian-2-yl)benzoate

To a dry vial was added the methyl 4-(1,3-dithian-2-yl)benzoate(KSC-335-053) (0.256 g, 1.01 mmol) and this was evacuated with argon 3times. The dry THF (7 mL) was added and the reaction was cooled to −78°C. at and the NaHMDS (1.258 mL, 1.258 mmol) was added. After 30 minutesthe 1-chloro-3-iodopropane (0.531 mL, 5.03 mmol) was added. The reactionwas then allowed to warm to rt overnight. The mixture was quenched withthe addition of saturated NH₄Cl (10 mL) at and diluted with EtOAc (15mL) and shaken. The EtOAc layer was collected, dried with MgSO₄,filtered and adsorbed to silica and purified by MPLC (20 min, −25%EtOAc:Hex) to produce pure methyl4-(2-(3-chloropropyl)-1,3-dithian-2-yl)benzoate (0.102 g, 0.308 mmol,31% yield).). ¹H NMR (400 MHz, CDCl₃): δ 8.07-8.03 (m, 2H), 8.01-7.98(m, 2H), 3.93 (s, 3H), 3.41 (t, J=6.4 Hz, 2H), 2.74-2.62 (m, 4H),2.19-2.13 (m, 2H), 1.99-1.92 (m, 2H), 1.78-1.70 (m, 2H).

KSC-335-059 (S)-1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-ol

To a flame-dried vial was added dry THF (2 mL) and then cooled to 0° C.The 1.0 M R-5,5-Diphenyl-2-methyl-3,4-propano-1,3,2-oxazaborlidine(0.105 mL, 0.105 mmol) in THF was added followed by the 2.0 Mborane-methyl sulfide complex (0.654 mL, 1.309 mmol) in THF. Thereaction began to stir at 0° C. for 30 minutes. To another flame-driedvial was added the 1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-one (0.250g, 1.047 mmol) and this was evacuated with argon 3 times then dissolvedin dry THF (5 mL) and the oxazaborlidine solution was added dropwise at0° C. and the reaction was allowed to warm to rt stirred for 2 h. Thereaction was quenched with MeOH (10 mL) extracted with EtOAc (20 mL)then was washed with 1.0 M HCl (3×25 mL). The EtOAc layer was dried withMgSO₄, filtered and concentrated to produce pure(S)-1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-ol (0.248 g, 1.030 mmol,98% yield). [alpha]²⁵ ₅₈₉=−24.0° (c=10 in CHCl₃). ¹H NMR (400 MHz,CDCl₃): δ 7.38 (d, J=8.4 Hz, 2H), 7.28 (d, J=8.4, 2H), 4.71-4.67 (m,1H), 3.61-3.53 (m, 2H), 1.98-1.79 (m, 4H), 1.32 (s, 9H).

KSC-335-060 (R)-1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-ol

Prepared the same as KSC-335-059 with 1.0 M(S)-5,5-diphenyl-2-methyl-3,4-propano-1,3,2-oxazaborlidine (0.105 ml,0.105 mmol), 2.0 M Borane-methyl sulfide complex (0.654 ml, 1.309 mmol)and 1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-one (0.250 g, 1.047 mmol)to produce pure (R)-1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-ol (0.216g, 0.897 mmol, 86% yield). [alpha]²⁵ ₅₈₉=+23.1° (c=10 in CHCl₃). ¹H NMR(400 MHz, CDCl₃): δ 7.38 (d, J=8.4 Hz, 2H), 7.28 (d, J=8.4, 2H),4.71-4.67 (m, 1H), 3.61-3.53 (m, 2H), 1.98-1.79 (m, 4H), 1.32 (s, 9H).

KSC-335-061 Methyl 4-(4-chlorobutanoyl)benzoate

To a vial was added the methyl4-(2-(3-chloropropyl)-1,3-dithian-2-yl)benzoate (KSC-335-056) (0.102 g,0.308 mmol) and acetonitrile (1.5 mL) with water (0.2 mL). The(bis(trifluoroacetoxy)iodo)benzene (0.199 g, 0.462 mmol) was then addedand the reaction stirred at rt for 1 h. The reaction was quenched withsaturated NaHCO₃ (7 mL) then diluted with EtOAc (10 mL) and extracted.The EtOAc was collected and washed with water (2×8 mL) and then driedwith MgSO₄, filtered and adsorbed to silica and purified by MPLC (20min, 0-25% EtOAc:hex to produce pure methyl 4-(4-chlorobutanoyl)benzoate(0.0512 g, 0.213 mmol, 69% yield). ¹H NMR (400 MHz, CDCl₃): δ 8.11 (d,J=8.00 Hz, 2H), 8.00 (d, J=8.6 Hz, 2H), 3.93 (s, 3H), 3.67 (t, J=6.1 Hz,2H), 3.19 (t, J=6.1 Hz, 2H), 2.22 (quintet, J=6.3 Hz, 2H).

KSC-335-062 (S)-1-(4-(tert-butyl)phenyl)-4-chlorobutyl acetate

To a vial was added the (S)-1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-ol(KSC-335-059) (0.248 g, 1.030 mmol) and diethyl ether (5.15 ml). The TEA(0.215 mL, 1.545 mmol) was added followed by the acetyl chloride (0.073mL, 1.030 mmol) and the reaction began to stir at rt for 2 h. A whiteprecipitate formed immediately. Water (5 mL) was added the reactionafter 2 h and the ether layer was extracted. The aqueous was extractedagain with more ether and the combined organics were dried with MgSO₄,filtered and concentrated to produce pure(S)-1-(4-(tert-butyl)phenyl)-4-chlorobutyl acetate (0.250 g, 0.884 mmol,86% yield). [alpha]²⁵ ₅₈₉=−42.11, (c=10, CH₂Cl₂). ¹H NMR (400 MHz,CDCl₃): δ 7.36 (d, J=8.4 Hz, 2H), 7.25 (d, J=8.4 Hz, 2H), 5.78-5.73 (m,1H), 3.53 (t, J=6.4 Hz, 2H), 2.06 (s, 3H), 2.02-1.68 (m, 4H), 1.31 (s,9H).

KSC-336-063 (R)-1-(4-(tert-butyl)phenyl)-4-chlorobutyl acetate

Prepared the same as KSC-335-062 with(R)-1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-ol (KSC-335-060) (0.216 g,0.897 mmol) and diethyl ether (4.5 mL), TEA (0.188 ml, 1.346 mmol) andacetyl chloride (0.064 ml, 0.897 mmol) to produce pure(R)-1-(4-(tert-butyl)phenyl)-4-chlorobutyl acetate (0.249 g, 0.880 mmol,98% yield). [alpha]²⁵ ₅₈₉=+54.57, (c=10, CH₂Cl₂). ¹H NMR (400 MHz,CDCl₃): δ 7.36 (d, J=8.4 Hz, 2H), 7.25 (d, J=8.4 Hz, 2H), 5.78-5.73 (m,1H), 3.53 (t, J=6.4 Hz, 2H), 2.06 (s, 3H), 2.02-1.68 (m, 4H), 1.31 (s,9H).

KSC-335-064(S)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butylacetate

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.154 g, 0.578mmol), (S)-1-(4-(tert-butyl)phenyl)-4-chlorobutyl acetate (0.196 g,0.693 mmol) (KSC-335-062) and potassium carbonate (0.319 g, 2.310 mmol)in acetonitrile (10 mL). The reaction stirred overnight at 70° C. for 18h and was then cooled to rt and filtered. The filtrate was then dilutedwith DCM and washed with water (10 mL) and brine (10 mL). The DCM layerswere combined, dried with MgSO₄, filtered and purified by reverse-phaseMPLC (20 min, 10-100% MeCN:water) to produce pure(S)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butylacetate (0.135 g, 0.263 mmol, 46% yield) as a brown oil. ¹H NMR (400MHz, CDCl₃): δ 7.48-7.45 (m, 4H), 7.35-7.14 (m, 10H), 5.73-5.68 (m, 1H),2.93-2.87 (m, 2H), 2.46-2.37 (m, 1H), 2.87 (t, J=7.7 Hz, 2H), 2.11 (brs, 1H), 2.04 (s, 3H), 1.95-1.84 (m, 3H), 1.80-1.72 (m, 1H), 1.53-1.38(m, 6H), 1.29 (s, 9H).

KSC-335-065(R)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butylacetate

Prepared the same way as KSC-335-064 withdiphenyl(piperidin-4-yl)methanol (0.196 g, 0.734 mmol),(R)-1-(4-(tert-butyl)phenyl)-4-chlorobutyl acetate (KSC-335-063) (0.249g, 0.880 mmol) and potassium carbonate (0.406 g, 2.93 mmol) inacetonitrile to produce pure(R)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butylacetate (0.237 g, 0.461 mmol, 63% yield) as a brown oil. ¹H NMR (400MHz, CDCl₃): δ 7.48-7.45 (m, 4H), 7.35-7.14 (m, 10H), 5.73-5.68 (m, 1H),2.93-2.87 (m, 2H), 2.46-2.37 (m, 1H), 2.87 (t, J=7.7 Hz, 2H), 2.11 (brs, 1H), 2.04 (s, 3H), 1.95-1.84 (m, 3H), 1.80-1.72 (m, 1H), 1.53-1.38(m, 6H), 1.29 (s, 9H).

KSC-335-066 Methyl2-(4-(4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butanoyl)phenyl)-2-methylpropanoate

To a vial was added the methyl2-(4-(4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)but-1-yn-1-yl)phenyl)-2-methylpropanoate(KSC-335-054) (0.074 g, 0.149 mmol). The mercuric oxide (1.493 ml, 0.045mmol) was made into a 0.03 M solution in 4% w/v sulfuric acid and addedto the starting material then heated to 55° C. and stirred for 3.5 h.The reaction turned a milky white color upon addition of the mercuricoxide solution. The reaction was removed from heat and diluted withsaturated NaHCO₃ (10 mL) and extracted with DCM (3×10 mL). The DCMlayers were combined and dried with MgSO₄, filtered and concentratedthen purified by reverse-phase MPLC (10-100% MeCN:water) to producemethyl 2-(4-(4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butanoyl)phenyl)-2-methylpropanoate (0.0217 g, 0.042mmol, 28.3% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.93-7.91 (m, 2H),7.48-7.45 (m, 4H), 7.42-7.39 (m, 2H), 7.30-7.26 (m, 4H), 7.19-7.14 (m,2H), 3.63 (s, 3H), 2.96-2.88 (m, 4H), 2.44-2.34 (m, 3H), 2.08 (br s,1H), 1.60 (s, 6H), 1.62-1.56 (m, 4H), 1.46-1.30 (m, 4H).

KSC-335-069(S)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol

To a vial was added the(S)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butylacetate (0.135 g, 0.263 mmol) and this vial was evacuated with nitrogen3 times. The dry THF (9 mL) was then added. The 1.0 M lithium aluminumhydride (0.263 mL, 0.263 mmol) in THF was added dropwise at rt and thereaction stirred for 5 h. The reaction was quenched slowly with water(10 mL) and then extracted with diethyl ether (2×10 mL). The ether layerwas dried with MgSO₄, filtered, concentrated and purified byreverse-phase MPLC (20 min, 10-100%, MeCN:H₂O) to produce pure(S)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol(0.100 g, 0.212 mmol, 81% yield). [alpha]²⁵ ₅₈₉=−38.8°. ¹H NMR (400 MHz,CDCl₃): δ 7.51-7.46 (m, 4H), 7.33-7.24 (m, 8H), 7.20-7.14 (m, 2H), 4.59(dd, J=8.2 Hz, 2.8 Hz, 1H), 3.13 (br d, J=11.2 Hz, 1H), 2.97 (br d,J=11.2 Hz, 1H), 2.50-2.34 (m, 3H), 2.10-1.88 (m, 4H), 1.84-1.74 (m, 1H),1.68-1.44 (m, 6H), 1.30 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ 149.4,146.1, 146.0, 142.7, 128.2, 128.1, 126.45, 126.43, 125.7, 125.6, 125.4,125.0, 79.3, 73.4, 58.9, 54.7, 53.3, 44.2, 39.7, 34.4, 31.4, 26.1, 26.0,24.1. LCMS Retention time: 4.159 min. LCMS purity 99.7%. HRMS (ESI): m/zcalcd for C₃₂H₄₁NO₂ [M+H]⁺ 472.3137. found 472.3210.

KSC-335-070(R)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol

To a vial was added the(R)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butylacetate (0.237 g, 0.461 mmol) and this vial was evacuated with nitrogen3 times. The dry THF (Volume: 4.61 ml) was added and then the 1.0 Mlithium aluminum hydride (0.461 mL, 0.461 mmol) in THF was addedportionwise at rt for 5 h. The reaction was quenched slowly with water(10 mL) and then extracted with diethyl ether (2×10 mL). The ether layerwas dried with MgSO₄, filtered, concentrated and purified byreverse-phase MPLC (20 min, 10-100%, MeCN:H₂O) to produce pure(R)-1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol (0.116 g, 0.246 mmol, 53% yield). [alpha]²⁵₅₈₉=+38.6 °. ¹H NMR (400 MHz, CDCl₃): δ 7.51-7.46 (m, 4H), 7.33-7.24 (m,8H), 7.20-7.14 (m, 2H), 4.59 (dd, J=8.2 Hz, 2.8 Hz, 1H), 3.13 (br d,J=11.2 Hz, 1H), 2.97 (br d, J=11.2 Hz, 1H), 2.50-2.34 (m, 3H), 2.10-1.88(m, 4H), 1.84-1.74 (m, 1H), 1.68-1.44 (m, 6H), 1.30 (s, 9H). ¹³C NMR(125 MHz, CDCl₃): δ 149.4, 146.1, 146.0, 142.7, 128.2, 128.1, 126.45,126.43, 125.7, 125.6, 125.4, 125.0, 79.3, 73.4, 58.9, 54.7, 53.3, 44.2,39.7, 34.4, 31.4, 26.1, 26.0, 24.1. LCMS Retention time: 4.156 min. LCMSpurity 97.8%. HRMS (ESI): m/z calcd for C₃₂H₄₁NO₂ [M+H]⁺ 472.3137. found472.3210.

KSC-335-0774-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(p-tolyl)butan-1-ol

Method B:4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(p-tolyl)butan-1-one(0.073 g, 0.171 mmol) and MeOH (Volume: 2 mL) and SODIUM BOROHYDRIDE(0.013 g, 0.341 mmol) to produce pure4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)-1-(p-tolyl)butan-1-ol (0.070g, 0.163 mmol, 95% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.51-7.46 (m, 4H), 7.32-7.09 (m, 10H), 4.59 (dd, J=8.0, 2.8 Hz, 1H),3.15-3.09 (m, 1H), 2.99-2.92 (m, 1H), 2.52-2.32 (m, 6H), 2.32 (s, 3H),2.09-1.87 (m, 3H), 1.82-1.73 (m, 1H), 1.68-1.44 (m, 6H). ¹³C NMR (125MHz, CDCl₃): δ 146.1, 146.0, 142.9, 136.0, 128.7, 128.11, 128.10,128.06, 126.39, 126.37, 125.7, 125.6, 125.5, 79.2, 73.3, 58.9, 54.6,53.4, 53.3, 44.2, 39.9, 30.9, 26.0, 25.9, 26.0, 21.0. LCMS Retentiontime: 3.809 min. LCMS purity 94.1%. HRMS (ESI): m/z calcd for C₂₉H₃₅NO₂[M+H]⁺ 430.2668. found 430.2741.

KSC-335-080 Methyl2-(4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)phenyl)-2-methylpropanoate

Method B: methyl 2-(4-(4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butanoyl)phenyl)-2-methylpropanoate (KSC-335-066)(0.148g, 0.288 mmol) and MeOH (1 mL) and sodium borohydride (0.016 g, 0.432mmol) to produce pure methyl2-(4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)phenyl)-2-methylpropanoate(0.108 g, 0.209 mmol, 73% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.54-7.49(m, 4H), 7.35-7.27 (m, 8H), 7.23-7.17 (m, 2H), 4.62 (dd, J=8.0, 2.8 Hz,1H), 3.65 (s, 3H), 3.16 (d, J=11.7 Hz, 1H), 2.98 (d, J=11.7 Hz, 1H),2.53-2.40 (m, 3H), 2.29 (br s, 1H), 2.14-2.06 (m, 1H), 2.01-1.93 (m,2H), 1.84-1.50 (m, 14H). LCMS Retention time: 3.786 min. LCMS purity98.2%. HRMS (ESI): m/z calcd for C₃₃H₄₁NO₄ [M+H]⁺ 516.3036. found516.3108.

KSC-335-0812-(4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)phenyl)-2-methylpropanoicacid

To a vial was added the methyl2-(4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)phenyl)-2-methylpropanoate(KSC-335-080) (0.094 g, 0.182 mmol) and THF (Volume: 3 mL,). The LiOH(0.022 g, 0.911 mmol) was dissolved in water (3.00 mL) and then added tothe reaction stirred at 80° C. for 18 h. The reaction was removed fromheat and cooled to rt and 1.0 M HCl in water was added to adjust to pHto 4 and a gummy off-white solid formed. DCM (5 mL) was added to themixture and it was sonicated to break up the solid. The DCM layer wasremoved and the aqueous was extracted with DCM (2×5 mL). The DCM layerwas concentrated and the residue was purified by reverse-phase MPLC (20min, 10-100% MeCN:water) to produce the product with impurities. Thiswas submitted to the purification core. The pure sample was recovered toproduce2-(4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)phenyl)-2-methylpropanoicacid (0.0378 g, 0.075 mmol, 41% yield). ¹H NMR (400 MHz, DMSO-d₆): δ8.22 (s, 1H), 7.52-7.49 (m, 4H), 7.29-7.26 (m, 8H), 7.15-7.10 (m, 2H),4.47 (t, J=5.9 Hz, 1H), 2.94-2.86 (m, 3H), 2.35-2.30 (m, 2H), 2.05-1.95(m, 2H), 1.60-1.35 (m, 12H), 1.28-1.22 (m, 2H). ¹³C NMR (125 MHz,CDCl₃): δ 177.7, 163.7, 147.2, 144.4, 143.4, 127.8, 125.8, 125.7, 125.6,125.1, 78.4, 71.8, 57.7, 53.4, 53.2, 45.5, 43.1, 37.3, 26.5, 25.6, 22.6.LCMS Retention time: 2.612 min. LCMS purity 100%. HRMS (ESI): m/z calcdfor C₃₂H₃₉NO₄ [M+H]⁺ 502.2879. found 502.2952.

KSC-342-0061-(4-(tert-butyl)phenyl)-2-(4-(hydroxydiphenylmethyl)piperidin-1-yl)ethanone

Method A: diphenyl(piperidin-4-yl)methanol (0.4 g, 1.496 mmol),1-(4-(tert-butyl)phenyl)-2-chloroethanone (0.300 g, 1.425 mmol), sodiumbicarbonate (0.144 g, 1.710 mmol) with water (3 mL) and 2-butanone(Volume: 15 mL) to produce pure1-(4-(tert-butyl)phenyl)-2-(4-(hydroxydiphenylmethyl)piperidin-1-yl)ethanone(0.452 g, 1.024 mmol, 71.8% yield) as a colorless oil. ¹H NMR (500 MHz,CDCl₃): δ 7.93 (d, J=8.6 Hz, 2H), 7.50-7.43 (m, 6H), 7.32-7.26 (m, 4H),7.20-7.15 (m, 2H), 3.78 (s, 2H), 3.07-3.01 (m, 2H), 2.47 (tt, J=11.8 Hz,3.5 Hz, 1H), 2.26-2.18 (m, 2H), 1.66-1.45 (m, 5H), 1.33 (s, 9H).). ¹³CNMR (125 MHz, CDCl₃): δ 196.1, 157.0, 145.8, 133.5, 129.8, 128.2, 128.0,126.5, 125.8, 125.5, 125.1, 79.5, 64.2, 54.2, 43.8, 35.1, 31.2, 31.0,26.2. LCMS Retention time: 3.987 min. LCMS purity 98.8%. HRMS (ESI): m/zcalcd for C₃₀H₃₅NO₂ [M+H]⁺ 442.2668. found 442.2741.

KSC-342-0101-(4-(tert-butyl)phenyl)-2-(4-(hydroxydiphenylmethyl)piperidin-1-yl)ethanol

Method B:1-(4-(tert-butyl)phenyl)-2-(4-(hydroxydiphenylmethyl)piperidin-1-yl)ethanone(0.113 g, 0.256 mmol) and MeOH (1 mL) and SODIUM BOROHYDRIDE (0.019 g,0.512 mmol) to produce pure1-(4-(tert-butyl)phenyl)-2-(4-(hydroxydiphenylmethyl)piperidin-1-yl)ethanol(0.106 g, 0.239 mmol, 93% yield) as a solid. ¹H NMR (400 MHz, CDCl₃): δ7.50-7.46 (m, 4H), 7.37-7.26 (m, 8H), 7.22-7.17 (M, 2H), 4.67 (m, 1H),4.01 (br s, 1H), 3.20 (m, 1H), 2.86 (m, 1H), 2.50-2.45 (m, 3H),2.37-2.30 (m, 1H), 2.10-2.02 (m, 1H), 1.60-1.45 (m, 5H), 1.31 (s, 9H).¹³C NMR (125 MHz, CDCl₃): δ 150.3, 145.82, 145.78, 139.1, 128.20,128.19, 126.59, 126.57, 125.7, 125.6, 125.2, 79.5, 68.6, 66.3, 55.8,53.4, 52.2, 44.1, 34.5, 31.3, 26.8, 26.5. LCMS Retention time: 4.083min. LCMS purity 94.2%. HRMS (ESI): m/z calcd for C₃₀H₃₇NO₂ [M+H]⁺444.2824. found 444.2897.

KSC-342-014 Methyl 4-(4-chloro-1-hydroxybutyl)benzoate

Method B: methyl 4-(4-chlorobutanoyl)benzoate (KSC-335-061) (0.042 g,0.175 mmol) and MeOH with sodium borohydride (0.013 g, 0.349 mmol). toproduce methyl 4-(4-chloro-1-hydroxybutyl)benzoate (0.037 g, 0.152 mmol,87% yield). ¹H NMR (400 MHz, CDCl₃): δ 8.00 (d, J=8.4 Hz, 2H), 7.40 (d,J=8.2 Hz, 2H), 4.8-4.6 (m, 1H), 3.90 (s, 3H), 3.58-3.52 (m, 2H), 2.19(br s, 1H), 1.96-1.78 (m, 4H)

KSC-342-017 Methyl4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)benzoate

To a vial was added the methyl 4-(4-chloro-1-hydroxybutyl)benzoate(KSC-342-014) (0.037 g, 0.152 mmol), diphenyl(piperidin-4-yl)methanol(0.122 g, 0.457 mmol), SODIUM BICARBONATE (0.026 g, 0.305 mmol), SODIUMIODIDE (1.143 mg, 7.62 μmol) and the vial was evacuated with argon 3times. Dry acetonitrile (2 ml) was added and the reaction stirredovernight at reflux and was then cooled to rt after 18 h and the solventwas concentrated. The residue was dissolved in DCM (5 mL) and washedwith 0.1 N HCl (5 mL), water (5 mL) and brine (5 mL). The product waspurified by MPLC (0-10% MeOH:DCM) to produce pure methyl4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)benzoate(0.0268 g, 0.057 mmol, 37% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.97 (d,J=8.4 Hz, 2H), 7.52-7.47 (m, 4H), 7.42 (d, J=7.9 Hz, 2H), 7.32-7.29 (m,4H), 7.20-7.14 (m, 2H), 4.66 (m, 1H), 3.90 (s, 3H), 3.14 (m, 1H), 2.94(m, 1H), 2.78 (br s, 1H), 2.52-2.43 (m, 1H), 2.39 (t, J=4.8 Hz, 2H),2.13-2.06 (m, 1H), 2.04-1.92 (m, 2H), 1.77-1.47 (m, 8H). ¹³C NMR (125MHz, CDCl₃): δ 167.2, 151.4, 146.0, 145.9, 129.5, 128.4, 128.2, 128.2,126.51, 126.48, 125.64, 125.59, 79.2, 73.2, 58.8, 54.7, 51.9, 44.2,40.0, 26.0, 25.9, 24.0. LCMS Retention time: 3.652 min. LCMS purity100%. HRMS (ESI): m/z calcd for C₃₀H₃₅NO₄ [M+H]⁺ 474.2566. found474.2639.

KSC-342-0214-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)benzoicacid

To a vial was added the methyl4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)benzoate(0.0195 g, 0.041 mmol) and THF (1 mL). The LiOH (6.90 mg, 0.288 mmol)was dissolved in water (1 mL) and added to the reaction. The reactionstirred at overnight and was then acidified with 1.0 M HCl to pH 2-3then extracted with DCM (3×5 mL). The DCM layer was concentrated andpurified by reverse-phase MPLC (10-100% MeCN:water) to produce pure4-(1-hydroxy-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butyl)benzoicacid (0.009 g, 0.020 mmol, 47% yield). ¹H NMR (400 MHz, CD₃OD): δ 7.85(d, J=8.2 Hz, 2H), 7.53-7.49 (m, 4H), 7.34-7.26 (m, 6H), 7.19-7.14 (m,2H), 4.70 (m, 1H), 3.46 (m, 1H), 3.35 (s, 2H), 3.01-2.96 (m, 2H),2.92-2.77 (m, 3H), 1.84-1.64 (m, 8H). ¹³C NMR (125 MHz, CD₃OD): δ 174.7,148.2, 147.2, 137.6, 130.4, 129.1, 127.6, 127.0, 126.4, 79.9, 74.0,53.9, 49.8, 36.9, 25.5, 21.8. LCMS Retention time: 2.490 min. LCMSpurity 100%. HRMS (ESI): m/z calcd for C₂₉H₃₃NO₄ [M+H]⁺ 460.2410. found460.2482.

KSC-342-074 1-(Tert-butyl)-4-(2-chloroethyl)benzene

To a vial was added the 1-(4-(tert-butyl)phenyl)-2-chloroethanone (0.171g, 0.812 mmol) and triethylsilane (0.519 mL, 3.25 mmol) with TFA (4 mL).The reaction stirred at 75° C. for 17 h and was then concentrated invacuo. The residue was dissolved in DCM (5 mL) and washed with water (4mL). The DCM layer was collected and washed with water (1×5 mL), driedwith MgSO₄, filtered and adsorbed to silica and purified by MPLC (20min, 0-40% EtOAc:hex) to produce pure1-(tert-butyl)-4-(2-chloroethyl)benzene (0.114 g, 0.580 mmol, 71%yield). ¹H NMR (400 MHz, CDCl₃): δ 7.34 (d, J=8.3 Hz, 2H), 7.16 (d,J=8.3 Hz, 2H), 3.71 (t, J=7.5 Hz, 2H), 3.05 (t, J=7.6 Hz, 2H), 1.32 (s,9H).

KSC-342-080(1-(3-(4-(Tert-butyl)phenyl)propyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.142 g, 0.530mmol), 1-(tert-butyl)-4-(3-chloropropyl)benzene (0.134 g, 0.636 mmol)and POTASSIUM CARBONATE (0.439 g, 3.18 mmol) in acetonitrile. Thereaction stirred overnight at 85° C. for 18 h and then filtered. Thefiltrate was then diluted with brine and extracted with diethyl ether(3×15 mL). The ether layers were combined, dried with MgSO₄, filteredand purified by reverse-phase MPLC (20 min, 10-100% MeCN:water) toproduce pure(1-(3-(4-(tert-butyl)phenyl)propyl)piperidin-4-yl)diphenylmethanol(0.152 g, 0.344 mmol, 65% yield) as an oil. ¹H NMR (400 MHz, CDCl₃): δ7.49-7.45 (m, 4H), 7.31-7.24 (m, 6H), 7.19-7.14 (m, 2H), 7.11-7.08 (m,2H), 2.96 (m, 2H), 2.57 (t, J=7.7 Hz, 2H), 2.48-2.32 (m, 3H), 2.22 (brs, 1H), 1.97-1.90 (m, 2H), 1.83-1.75 (m, 2H), 1.53-1.45 (m, 4H), 1.29(s, 9H). LCMS Retention time: 2.965 min. LCMS purity 97.2%. HRMS (ESI):m/z calcd for C₃₁H₃₉NO [M+H]⁺ 442.3032. found 442.3104.

KSC-342-081 (1-(4-(Tert-butyl)phenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.150 g, 0.561mmol), 1-(tert-butyl)-4-(2-chloroethyl)benzene (KSC-342-074) (0.110 g,0.561 mmol) and potassium carbonate (0.465 g, 3.37 mmol) inacetonitrile. The reaction stirred overnight at 85° C. for 18 h and thenfiltered. The filtrate was then diluted with brine and extracted withdiethyl ether (3×15 mL). The ether layers were combined, dried withMgSO₄, filtered and purified by reverse-phase MPLC (20 min, 10-100%MeCN:water) to produce pure(1-(4-(tert-butyl)phenethyl)piperidin-4-yl)diphenylmethanol (0.180 g,0.421 mmol, 75% yield) as an oil. ¹H NMR (400 MHz, CDCl₃): δ 7.51-7.47(m, 4H), 7.33-7.27 (m, 6H), 7.21-7.16 (m, 2H), 7.14-7.11 (m, 2H),3.09-3.03 (m, 2H), 2.80-2.74 (m, 2H), 2.60-2.55 (m, 2H), 2.51-2.42 (m,1H), 2.28 (br s, 1H), 2.08-2.00 (m, 2H), 1.57-1.50 (m, 4H), 1.30 (s,9H). ¹³C NMR (125 MHz, CDCl₃): δ 148.8, 145.9, 137.3, 128.3, 128.1,126.5, 125.8, 125.2, 79.5, 60.8, 54.0, 44.2, 40.9, 34.3, 33.1, 31.4,26.4. LCMS Retention time: 4.384 min. LCMS purity 99.7%. HRMS (ESI): m/zcalcd for C₃₀H₃₇NO [M+H]⁺ 428.2875. found 428.2948.

KSC-342-088(1-(4-amino-4-(4-(tert-butyl)phenyl)butyl)piperidin-4-yl)diphenylmethanol

To a vial was added the1-(4-(tert-butyl)phenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(0.200 g, 0.426 mmol), Ammonium acetate (0.328 g, 4.26 mmol) and Sodiumcyanoborohydride (0.040 g, 0.639 mmol) with MeOH (Volume: 4 ml). Thereaction stirred at rt at 2:56:22 PM. The reaction was stirred overnightand then concentrated and diluted with dilute aqueous ammonium hydroxideand extracted with DCM. The DCM layer was concentrated and the crude NMRshowed product and starting material. The reaction then purified byreverse-phase Teledyne ISCO Combiflash chromatography (10-100%MeCN:basic water) and fractions 5 and 6 were collected. These were thensubjected to normal phase purification (0-10% MeOH (5% NH₃OH):DCM) toproduce pure(1-(4-amino-4-(4-(tert-butyl)phenyl)butyl)piperidin-4-yl)diphenylmethanol(0.010 g, 0.021 mmol, 5% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.48-7.44(m, 4H), 7.34-7.26 (m, 6H), 7.22-7.14 (m, 4H), 3.84 (t, J=6.7 Hz, 1H),2.95-2.87 (m, 2H), 2.46-2.37 (m, 1H), 2.28 (t, J=7.6 Hz, 2H), 1.95-1.85(m, 2H), 1.70-1.60 (m, 5H), 1.55-1.40 (m, 6H), 1.30 (s, 9H). ¹³C NMR(125 MHz, CDCl₃): δ 149.7, 146.0, 143.3, 128.1, 126.5, 125.9, 125.8,125.3, 79.5, 58.7, 55.8, 54.2, 54.0, 44.2, 37.5, 34.4, 31.4, 26.4, 24.2.LCMS Retention time: 2.397 min. LCMS purity 100%. HRMS (ESI): m/z calcdfor C₃₂H₄₂N₂O [M+H]⁺ 471.3297. found 471.3370.

KSC-348-0011-([1,1′-Biphenyl]-4-yl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one

To a vial was added the1-(4-bromophenyl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(0.098 g, 0.199 mmol), 1,1′-bis(di-t-butylphosphino)ferrocene palladiumdichloride, (2.71 mg, 3.98 μmol) and phenylboronic acid (0.029 g, 0.239mmol) followed by acetonitrile (1.5 mL). The potassium carbonate (0.041g, 0.299 mmol) was dissolved in water (1.5 mL) and added the reaction.The reaction stirred at 60° C. for 18 h. The reaction was stopped andthe organic layer was diluted with EtOAc and extracted then washed withbrine. The EtOAc layer was collected, dried with MgSO₄, filtered andpurified by reverse-phase MPLC (10-100% MeCN:water) to produce thedesired1-([1,1′-biphenyl]-4-yl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(0.035 g, 0.071 mmol, 36% yield). ¹H NMR (400 MHz, CDCl₃): δ 8.03 (d,J=8.6 Hz, 2H), 7.67 (d, J=8.4 Hz, 2H), 7.64-7.61 (m, 2H), 7.50-7.45 (m,6H), 7.42-7.38 (m, 1H), 7.31-7.26 (m, 4H), 7.19-7.14 (m, 2H), 3.02-2.91(m, 4H), 2.46-2.37 (m, 3H), 2.10 (br s, 1H), 1.97-1.92 (m, 4H),1.50-1.35 (m, 4H).

KSC-348-0021-([1,1′-biphenyl]-4-yl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol

Method B:1-([1,1′-biphenyl]-4-yl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-one(KSC-348-001) (0.035 g, 0.071 mmol) and MeOH (2 mL) and sodiumborohydride (5.41 mg, 0.143 mmol) to produce pure1-([1,1′-biphenyl]-4-yl)-4-(4-(hydroxydiphenylmethyl)piperidin-1-yl)butan-1-ol(0.033 g, 0.067 mmol, 94% yield) as on oil. ¹H NMR (400 MHz, CDCl₃): δ7.59-7.55 (m, 2H), 7.53-7.46 (m, 6H), 7.43-7.38 (m, 4H), 7.34-7.25 (m,5H), 7.19-7.13 (m, 2H), 4.65 (dd, J=8.2 Hz, 2.7 Hz, 1H), 3.16-3.10 (m,1H), 2.99-2.93 (m, 1H), 2.56 (br s, 1H), 2.50-2.34 (m, 3H), 2.10-1.92(m, 3H), 1.86-1.76 (m, 1H), 1.70-1.45 (m, 6H).). ¹³C NMR (125 MHz,CDCl₃): δ 146.1, 146.0, 145.0, 141.1, 139.4, 128.6, 128.2, 128.1,127.01, 126.96, 126.8, 126.44, 126.41, 126.1, 125.7, 125.6, 79.2, 73.3,58.9, 54.7, 53.3, 44.2, 40.0, 26.0, 25.9, 24.1. LCMS Retention time:4.049 min. LCMS purity 97.6%. HRMS (ESI): m/z calcd for C₃₄H₃₇NO₂ [M+H]⁺492.2824. found 492.2897.

KSC-348-049(4-(tert-butyl)phenyl)(4-(hydroxydiphenylmethyl)piperidin-1-yl)methanone

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.232 g, 0.868mmol), acetonitrile (3 mL) and TEA (0.181 ml, 1.30 mmol). The4-(tert-butyl)benzoyl chloride (0.173 mL, 0.954 mmol) was added and thereaction stirred at 70° C. for 6 h and was diluted with EtOAc (15 mL)and washed with saturated NaHCO₃ (15 mL). The EtOAc was collected, driedwith MgSO₄, filtered and adsorbed to silica and purified by MPLC (20min, 0-30% EtOAc:Hex) to produce pure(4-(tert-butyl)phenyl)(4-(hydroxydiphenylmethyl)piperidin-1-yl)methanone(0.304 g, 0.711 mmol, 82% yield) as a white solid. ¹H NMR (400 MHz,CDCl₃): δ 7.50-7.44 (m, 4H), 7.38-7.14 (m, 10H), 4.77 (br s, 1H), 3.84(br s, 1H), 3.05-2.63 (m, 3H), 2.25-2.17 (m, 1H), 1.75-1.35 (m, 4H),1.29 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ 170.4, 152.7, 145.4, 133.2,128.3, 126.7, 125.7, 125.2, 79.5, 44.5, 34.7, 31.2. LCMS Retention time:3.774 min. LCMS purity 100%. HRMS (ESI): m/z calcd for C₂₉H₃₃NO₂ [M+H]⁺428.2511. found 428.2584.

KSC-348-050 (1-(4-(tert-butyl)benzyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.085 ml,0.393 mmol), acetonitrile (2 mL) and TEA (0.082 ml, 0.589 mmol). Thep-tert-butylbenzyl bromide (0.098 g, 0.432 mmol) was then added and thereaction stirred at 70° C. and stirred for 5 h then diluted with EtOAc(15 mL) and washed with saturated NaHCO₃ (15 mL). The EtOAc wascollected, dried with MgSO₄, filtered and adsorbed to silica andpurified by reverse phase MPLC (20 min, 10-100% MeCN:water) to producepure (1-(4-(tert-butyl)benzyl)piperidin-4-yl)diphenylmethanol (0.131 g,0.317 mmol, 81% yield) as a brown oil. ¹H NMR (400 MHz, CDCl₃): δ7.48-7.45 (m, 4H), 7.32-7.25 (m, 6H), 7.22-7.14 (m, 4H), 3.47 (s, 2H),2.96-2.90 (m, 2H), 2.46-2.38 (m, 1H), 2.02-1.95 (m, 1H), 1.51-1.44 (m,4H), 1.30 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ 149.8, 146.0, 135.1,128.9, 128.1, 126.4, 125.8, 125.0, 79.5, 62.8, 53.9, 44.2, 34.4, 31.4,26.5, 21.0. LCMS Retention time: 4.186 min. LCMS purity 99.7%. HRMS(ESI): m/z calcd for C₂₉H₃₅NO [M+H]⁺ 414.2719. found 414.2791.

KSC-348-058 (1-(4-methoxyphenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.381 g, 1.425mmol), acetonitrile (5 mL) and TEA (0.298 ml, 2.138 mmol). The1-(2-bromoethyl)-4-methoxybenzene (0.245 ml, 1.568 mmol) was then addedand the reaction stirred at 75° C. and the stirred for 16 h then wasquenched with saturated NaHCO₃, extracted with EtOAc, dried with MgSO₄,filtered and adsorbed to silica. The product was purified by MPLC (20min, 0-10% MeOH:DCM) to produce pure (1-(4methoxyphenethyl)piperidin-4-yl)diphenylmethanol (0.289 g, 0.720 mmol, 50% yield) as asticky solid. ¹H NMR (400 MHz, CDCl₃): δ 7.49-7.46 (m, 4H), 7.32-7.28(m, 4H), 7.19 (tt, J=7.3 Hz, 1.9 Hz, 2H), 7.12 (d, J=8.6 Hz, 2H), 6.82(d, J=8.5 Hz, 2H), 3.77 (s, 3H), 3.29-3.24 (m, 2H), 2.95-2.90 (m, 2H),2.80-2.75 (m, 2H), 2.58-2.50 (m, 1H), 2.42-2.30 (m, 3H), 1.90-1.77 (m,2H), 1.65-1.57 (m, 2H). ¹³C NMR (125 MHz, CDCl₃): δ 158.3, 145.5, 129.6,128.3, 126.7, 125.6, 114.0, 79.3, 55.3, 53.6, 53.4, 43.4, 31.5, 25.1.LCMS Retention time: 3.695 min. LCMS purity 100%. HRMS (ESI): m/z calcdfor C₂₇H₃₁NO₂ [M+H]⁺ 402.2355. found 402.2353.

KSC-352-0554-(4-Benzoylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-one

To a vial was added the 1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-one(0.060 g, 0.252 mmol) and potassium iodide (0.063 g, 0.377 mmol) withacetonitrile (2 mL). The reaction stirred at 85° C. for 1 h then thephenyl(piperidin-4-yl)methanone (0.050 g, 0.264 mmol) along withpotassium carbonate (0.052 g, 0.377 mmol) was added. The reaction wasthen heated back to 85° C. for 48 h. The reaction was cooled to rt anddiluted with water then extracted with EtOAc (3×15 mL). The EtOAc layerwas dried with MgSO₄, filtered and adsorbed to silica. The product waspurified by reverse-phase MPLC (20 min, 10-100% MeCN:water) to produce4-(4-benzoylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-one (0.026g, 0.066 mmol, 26% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.95-7.90 (m, 4H),7.57-7.53 (m, 1H), 7.49-7.44 (m, 4H), 3.27-3.18 (m, 1H), 3.03-2.96 (m,4H), 2.44 (t, J=7.04 Hz, 2H), 2.13-2.06 (m, 2H), 1.98-1.91 (m,

2H), 1.85-1.76 (m, 4H), 1.34 (s, 9H). KSC-352-060(1-(4-nitrophenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.513 g, 1.919mmol), 1-(2-bromoethyl)-4-nitrobenzene (0.401 g, 1.744 mmol) andacetonitrile (10 mL). The TEA (0.365 ml, 2.62 mmol) was then added andthe reaction stirred at 85° C. for 18 h then cooled to rt. The reactionwas diluted with water and extracted with EtOAc (3×15 mL). The EtOAclayer was collected and dried with MgSO₄, filtered and adsorbed tosilica then purified by MPLC (15 min, 0-10% MeOH:DCM). to produce pure(1-(4-nitrophenethyl)piperidin-4-yl)diphenylmethanol (0.145 g, 0.348mmol, 20% yield). ¹H NMR (400 MHz, CDCl₃): δ 8.13 (d, J=8.7 Hz, 2H),7.49-7.46 (m, 4H), 7.35 (d, J=8.7 Hz, 2H), 7.32-7.27 (m, 4H), 7.21-7.16(m, 2H), 3.11-3.05 (m, 2H), 2.97-2.91 (m, 2H), 2.69-2.63 (m, 2H),2.52-2.43 (m, 1H), 2.25 (br s, 1H), 2.20-2.10 (m, 2H), 1.63-1.53 (m,4H). ¹³C NMR (125 MHz, CDCl₃): δ 147.9, 146.5, 145.7, 129.5, 128.2,126.6, 125.7, 123.7, 79.4, 59.4, 53.9, 43.9, 33.2, 26.1, 21.1. LCMSRetention time: 3.654 min. LCMS purity 98.8%. HRMS (ESI): m/z calcd forC₂₆H₂₈N₂O₃ [M+H]⁺ 417.2162. found 417.2173.

KSC-352-061 (1-(4-bromophenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.496 g, 1.855mmol), 1-bromo-4-(2-bromoethyl)benzene (0.258 ml, 1.69 mmol) andacetonitrile (10 mL). The TEA (0.353 mL, 2.53 mmol) was then added andthe reaction stirred at 85° C. for 18 h then cooled to rt. The reactionwas diluted with water and extracted with EtOAc (3×15 mL). The EtOAclayer was collected and dried with MgSO₄, filtered and adsorbed tosilica then purified by MPLC (15 min, 0-10% MeOH:DCM). to produce pure(1-(4-bromophenethyl)piperidin-4-yl)diphenylmethanol (0.488 g, 1.083mmol, 64% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.50-7.46 (m, 4H), 7.38 (d,J=8.3 Hz, 2H), 7.34-7.27 (m, 4H), 7.18 (tt, J=7.3 Hz, 1.8 Hz, 2H), 7.06(d, J=8.4 Hz, 2H), 3.06-3.00 (m, 2H), 2.77-2.72 (m, 2H), 2.58-2.52 (m,2H), 2.50-2.41 (m, 1H), 2.15-2.00 (m, 3H), 1.56-1.49 (m, 4H). ¹³C NMR(125 MHz, CDCl₃): δ 145.8, 139.3, 131.4, 130.4, 128.2, 126.6, 125.8,119.8, 79.5, 60.4, 54.0, 44.1, 33.1, 26.4, 21.1 LCMS Retention time:3.969 min. LCMS purity 99.8%. HRMS (ESI): m/z calcd for C₂₆H₂₈BrNO[M+H]⁺ 450.1354. found 450.1427.

KSC-352-063(1-(2-([1,1′-biphenyl]-4-yl)ethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the phenylboronic acid (0.019 g, 0.152 mmol),1,1′-bis(di-tert-butylphosphino)ferrocene palladium dichloride (4.12 mg,6.33 μmol), (1-(4-bromophenethyl)piperidin-4-yl)diphenylmethanol(KSC-352-061) (0.057 g, 0.127 mmol) and potassium carbonate (0.035 g,0.253 mmol). The vial was then evacuated with argon 3 times andacetonitrile (1 mL) was added followed by water (1 mL). The reactionthen stirred at 60° C. for 18 h then was diluted with EtOAc (5 mL) andsaturated NaHCO₃ (5 mL). The EtOAc layer was collected and the aqueouslayer was extracted with more EtOAc (2×5 mL). The EtOAc layers werecombined and dried with MgSO₄, filtered and concentrated. The reactionwas purified by MPLC (10-100% MeCN:water) to produce pure(1-(2-([1,1′-biphenyl]-4-yl)ethyl)piperidin-4-yl)diphenylmethanol (0.045g, 0.101 mmol, 79% yield) as a clear oil. ¹H NMR (400 MHz, CDCl₃): δ7.59-7.45 (m, 2H), 7.53-7.48 (m, 6H), 7.45-7.40 (m, 2H), 7.35-7.25 (m,7H), 7.22-7.16 (m, 2H), 3.11-3.05 (m, 2H), 2.87-2.81 (m, 2H), 2.65-2.59(m, 2H), 2.53-2.44 (m, 1H), 2.22 (br s, 1H), 2.12-2.04 (m, 2H),1.59-1.52 (m, 4H). ¹³C NMR (125 MHz, CDCl₃): δ 145.9, 141.0, 139.5,139.0, 129.1, 128.7, 128.2, 127.1, 127.03, 126.96, 126.5, 125.8, 79.5,60.7, 54.0, 50.8, 44.2, 33.3, 26.4. LCMS Retention time: 4.086 min. LCMSpurity 99%. HRMS (ESI): m/z calcd for C₃₂H₃₃NO [M+H]⁺ 448.2562. found448.2635.

KSC-352-064Diphenyl(1-(4-(pyridin-4-yl)phenethyl)piperidin-4-yl)methanol

To a vial was added the(1-(4-bromophenethyl)piperidin-4-yl)diphenylmethanol (KSC-352-061)(0.050 g, 0.111 mmol), pyridin-4-ylboronic acid (0.018 g, 0.133 mmol),1,1′-bis(di-tert-butylphosphino)ferrocene palladium dichloride (3.62 mg,5.55 μmol) and potassium carbonate (0.031 g, 0.222 mmol). The vial wasevacuated 3 times with argon and then acetonitrile (1 mL) followed bywater (1 mL) was added. The reaction stirred at 60° C. for 18 h and wasthen cooled to rt and diluted with EtOAc (5 mL) and saturated NaHCO₃ (5mL). The EtOAc layer was collected and the aqueous layer was extractedwith more EtOAc (2×5 mL). The EtOAc layers were combined and dried withMgSO₄, filtered and concentrated. The reaction was purified byreverse-phase MPLC (10-100% MeCN:water) to produce purediphenyl(1-(4-(pyridin-4-yl)phenethyl)piperidin-4-yl)methanol (0.015 g,0.033 mmol, 30% yield) as an oil. ¹H NMR (400 MHz, CDCl₃): δ 8.64-8.62(m, 2H), 7.60 (d, J=8.2 Hz, 2H), 7.51-7.46 (m, 6H), 7.32-7.28 (m, 6H),7.18 (tt, J=7.3 Hz, 1.8 Hz, 2H), 3.10-3.04 (m, 2H), 2.88-2.82 (m, 2H),2.65-2.58 (m, 2H), 2.51-2.43 (m, 1H), 2.19 (br s, 1H), 2.12-2.03 (m,2H), 1.58-1.51 (m, 4H). ¹³C NMR (125 MHz, CDCl₃): δ 150.2, 148.1, 145.9,141.7, 135.8, 129.5, 128.2, 127.0, 126.5, 125.8, 121.4, 79.5, 60.5,54.1, 44.1, 41.0, 33.4, 26.4. LCMS Retention time: 3.585 min. LCMSpurity 93.3%. HRMS (ESI): m/z calcd for C₃₁H₃₂N₂O [M+H]⁺ 449.2515. found449.2587.

KSC-352-065Diphenyl(1-(4-(pyridin-3-yl)phenethyl)piperidin-4-yl)methanol

To a vial was added the(1-(4-bromophenethyl)piperidin-4-yl)diphenylmethanol (0.057 g, 0.127mmol), pyridin-3-ylboronic acid (0.019 g, 0.152 mmol),1,1′-Bis(di-tert-butylphosphino)ferrocene palladium dichloride (4.12 mg,6.33 μmol) and potassium carbonate (0.035 g, 0.253 mmol). The vial wasevacuated with argon 3 times and then acetonitrile (1 mL) followed bywater (1 mL) was added. The reaction stirred at 60° C. for 18 h and thencooled to rt and diluted with EtOAc (5 mL) and saturated NaHCO₃ (5 mL).The EtOAc layer was collected and the aqueous layer was extracted withmore EtOAc (2×5 mL). The EtOAc layers were combined and dried withMgSO₄, filtered and concentrated. The reaction was purified by RP MPLC(10-100% MeCN:water) to produce purediphenyl(1-(4-(pyridin-3-yl)phenethyl)piperidin-4-yl)methanol (0.05 g,0.111 mmol, 88% yield) as an oil. ¹H NMR (400 MHz, CDCl₃): δ 8.79 (dd,J=2.4 Hz, 0.9 Hz, 1H), 8.53 (dd, J=4.8 Hz, 1.6 Hz, 1H), 7.86-7.83 (m,1H), 7.51-7.47 (m, 6H), 7.36-7.27 (m, 7H), 7.20-7.15 (m, 2H), 3.10-3.03(m, 2H), 2.87-2.81 (m, 2H), 2.71 (br s, 1H), 2.63-2.58 (m, 2H),2.51-2.43 (m, 1H), 2.11-2.03 (m, 2H), 1.58-1.52 (m, 4H).). ¹³C NMR (125MHz, CDCl₃): δ 148.04, 147.97, 146.0, 140.5, 136.5, 135.5, 134.3, 129.4,128.1, 127.1, 126.4, 125.8, 123.5, 79.4, 60.5, 54.1, 44.1, 33.2, 26.2.LCMS Retention time: 3.582 min. LCMS purity 98.8%. HRMS (ESI): m/z calcdfor C₃₁H₃₂N₂O [M+H]⁺ 449.2515. found 449.2587.

KSC-352-0661-(4-(tert-butyl)phenyl)-4-(4-(hydroxy(phenyl)methyl)piperidin-1-yl)butan-1-ol

Method B:4-(4-benzoylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-one(KSC-352-055) (0.026 g, 0.066 mmol) and MeOH (2 mL) and sodiumborohydride (10.05 mg, 0.266 mmol) to produce pure1-(4-(tert-butyl)phenyl)-4-(4-(hydroxy(phenyl)methyl)piperidin-1-yl)butan-1-ol(0.019 g, 0.048 mmol, 72% yield) as a mixture of diastereomers. ¹H NMR(400 MHz, CDCl₃): δ 7.36-7.24 (m, 9H), 4.63-4.57 (m, 1H), 4.33 (d, J=7.6Hz, 1H), 3.23-2.83 (m, 2H), 2.42-2.35 (m, 2H), 2.13-1.89 (m, 3H),1.86-1.52 (m, 6H), 1.46-1.16 (m, 4H), 1.31 (s, 9H). ¹³C NMR (125 MHz,CDCl₃): δ 149.4, 143.4, 142.8, 128.3, 127.63, 127.61, 126.6, 125.4,125.0, 78.74, 78.67, 73.4, 58.9, 54.30, 54.26, 52.8, 52.7, 43.21, 43.2,39.9, 34.4, 31.4, 28.4, 28.3, 28.2, 24.2, 24.1. LCMS Retention time:3.707 min. LCMS purity 97.1%. HRMS (ESI): m/z calcd for C₂₆H₃₇NO₂ [M+H]⁺396.2824. found 396.2897.

KSC-352-069 (1-(4-aminophenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the (1-(4-nitrophenethyl)piperidin-4-yl)diphenylmethanol (KSC-352-060) (0.135 g, 0.324 mmol) withMeOH (1 mL) and DCM (1 mL). The reaction was cooled to 0° C. and theRaney Nickel (1.902 mg, 0.032 mmol) was added. The sodium borohydride(0.031 g, 0.810 mmol) was then added portionwise and the reactionstirred at rt for 28 h and the Raney Nickel filtered through celite. Thereaction was diluted with DCM and washed with water and the DCM layerwas dried with MgSO₄, filtered and adsorbed to silica then purified byMPLC (0-15% MeOH:DCM) to produce pure(1-(4-aminophenethyl)piperidin-4-yl)diphenylmethanol (0.077 g, 0.199mmol, 61% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.51-7.47 (m, 4H),7.32-7.27 (m, 4H), 7.18 (tt, J=7.3 Hz, 1.8 Hz, 2H), 6.97 (d, J=8.3 Hz,2H), 6.61 (d, J=8.3 Hz, 2H), 3.52 (br s, 2H), 3.07-3.01 (m, 2H),2.70-2.65 (m, 2H), 2.54-2.41 (m, 3H), 2.22 (br s, 1H), 2.07-1.98 (m,2H), 1.56-1.49 (m, 4H). ¹³C NMR (125 MHz, CDCl₃): δ 145.9, 144.4, 130.4,129.4, 128.1, 126.5, 125.8, 115.2, 79.5, 61.2, 54.1, 44.2, 32.8, 26.4,21.0. LCMS Retention time: 3.328 min. LCMS purity 93.1%. HRMS (ESI): m/zcalcd for C₂₆H₃₀N₂O [M+H]⁺ 387.2358. found 387.2431.

KSC-352-075(1-(4-(Dimethylamino)phenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the(1-(4-aminophenethyl)piperidin-4-yl)diphenylmethanol (KSC-352-069)(0.030 g, 0.078 mmol) and acetic acid (1 mL). The paraformaldehyde(0.058 mL, 0.776 mmol) solution in water followed by sodiumcyanoborohydride (0.015 g, 0.233 mmol) was then added and the reactionstirred at rt for 20 h. The reaction was concentrated and diluted withsaturated NaHCO₃ and extracted with EtOAc (3×5 mL). The EtOAc layer wasdried with MgSO₄, filtered and concentrated. The product was purified byreverse-phase MPLC (10-100% MeCN:water) to produce(1-(4-(dimethylamino)phenethyl)piperidin-4-yl)diphenylmethanol (0.027 g,0.065 mmol, 84% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.50-7.46 (m, 4H),7.31-7.26 (m, 4H), 7.17 (tt, J=7.3 Hz, 1.8 Hz, 2H), 7.06 (d, J=8.3 Hz,2H), 6.68 (d, J=8.3 Hz, 2H), 3.08-3.02 (m, 2H), 2.90 (s, 6H), 2.72-2.67(m, 2H), 2.56-2.50 (m, 2H), 2.48-2.42 (m, 1H), 2.30 (br s, 1H),2.07-2.00 (m, 2H), 1.56-1.51 (m, 4H). ¹³C NMR (125 MHz, CDCl₃): δ 149.1,146.0, 129.2, 128.4, 128.1, 126.4, 125.8, 112.9, 79.4, 61.1, 54.0, 44.2,41.0, 40.8, 32.6, 26.4. LCMS Retention time: 2.50 min. LCMS purity96.1%. HRMS (ESI): m/z calcd for C₂₈H₃₄N₂O [M+H]⁺ 415.2671. found415.2744.

KSC-352-082Diphenyl(1-(4-(trifluoromethyl)phenethyl)piperidin-4-yl)methanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.100 g, 0.374mmol), 1-(2-bromoethyl)-4-(trifluoromethyl)benzene (0.057 ml, 0.340mmol) and acetonitrile (10 mL). The TEA (0.071 mL, 0.510 mmol) was thenadded and the reaction stirred at 85° C. for 18 h. The reaction wasdiluted with water and extracted with EtOAc (3×15 mL). The EtOAc layerwas collected and dried with MgSO₄, filtered and adsorbed to silica thenpurified by MPLC (15 min, 0-10% MeOH:DCM) to produce purediphenyl(1-(4-(trifluoromethyl)phenethyl)piperidin-4-yl)methanol (0.074g, 0.168 mmol, 49% yield) as an oil. ¹H NMR (400 MHz, CDCl₃): δ7.54-7.47 (m, 4H), 7.33-7.27 (m, 6H), 7.19 (tt, J=7.3 Hz, 1.8 Hz, 2H),3.06-3.00 (m, 2H), 2.86-2.80 (m, 2H), 2.60-2.55 (m, 2H), 2.51-2.42 (m,1H), 2.14 (br s, 1H), 2.10-2.03 (m, 2H), 1.57-1.47 (m, 4H). ¹³C NMR (125MHz, CDCl₃): δ 145.9, 144.6, 129.0, 128.3 (q, J=32 Hz), 128.2, 126.5,125.8, 125.2 (q, 3.8 Hz), 124.2 (q, J=271.8 Hz), 79.5, 60.2, 54.1, 44.1,33.6, 26.4, 21.0. LCMS Retention time: 3.877 min. LCMS purity 99.1%.HRMS (ESI): m/z calcd for C₂₇H₂₈F₃NO [M+H]⁺ 440.2123. found 440.2196.

KSC-352-088 (1-(4-Fluorophenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.099 g, 0.370mmol), 1-(2-bromoethyl)-4-fluorobenzene (0.047 mL, 0.337 mmol) andacetonitrile (10 mL). The TEA (0.070 mL, 0.505 mmol) was then added andthe reaction stirred at 85° C. at for 19 h and was cooled to rt. Thereaction was diluted with water and extracted with EtOAc (3×15 mL). TheEtOAc layer was collected and dried with MgSO₄, filtered and adsorbed tosilica then purified by MPLC (15 min, 0-10% MeOH:DCM) to produce pure(1-(4-fluorophenethyl)piperidin-4-yl)diphenylmethanol (0.123 g, 0.316mmol, 94% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.51-7.47 (m, 4H),7.32-7.27 (m, 4H), 7.21-7.11 (m, 4H), 6.98-6.92 (m, 2H), 3.06-3.00 (m,2H), 2.78-2.72 (m, 2H), 2.56-2.51 (m, 2H), 2.50-2.42 (m, 1H), 2.21 (brs, 1H), 2.08-1.99 (m, 2H), 1.57-1.48 (m, 4H). ¹³C NMR (125 MHz, CDCl₃):δ 161.2 (d, J=243.6 Hz), 145.9, 136.5 (d, J=3.2 Hz), 129.9 (d, J=7.9Hz), 128.1, 126.5, 125.8, 115.0 (d, J=21.2 Hz), 79.5, 60.8, 54.1, 44.2,33.0, 26.4, 21.0. LCMS Retention time: 3.733 min. LCMS purity 96.8%.HRMS (ESI): m/z calcd for C₂₆H₂₈FNO [M+H]⁺ 390.2155. found 390.2228.

KSC-352-090 2-(3-(Tert-butyl)phenyl)ethanol

To a vial was added the 1-bromo-3-(tert-butyl)benzene (0.107 g, 0.502mmol) and dry THF. The reaction was then cooled to −78° C. and the BuLi(0.221 mL, 0.552 mmol) (2.5 M in hexanes) was added dropwise and thereaction stirred for 30 min at −78° C. and then ethylene oxide (0.502mL, 1.255 mmol) (2.5-3.3M solution in THF) was added dropwise and thereaction stirred for 10 minutes at −78° C. then warmed to rt and stirredfor 1 h. The reaction was then quenched with 1.0 M HCl (2 mL) andextracted with EtOAc (3×5 mL). The EtOAc layer was combined,concentrated and purified by reverse-phase MPLC (10-100% MeCN:water) toproduce pure 2-(3-(tert-butyl)phenyl)ethanol (0.035 g, 0.196 mmol, 39%yield). ¹H NMR (400 MHz, CDCl₃): δ 7.31-7.27 (m, 3H), 7.10-7.07 (m, 1H),3.90 (t, J=6.6 Hz, 2H), 2.91 (t, J=6.5 Hz, 2H), 1.52 (br s, 1H), 1.36(s, 9H).

KSC-352-093 3-(Tert-butyl)phenethyl 4-methylbenzenesulfonate

To a vial was added the 2-(3-(tert-butyl)phenyl)ethanol (KSC-352-090)(0.035 g, 0.196 mmol), TEA (0.082 mL, 0.589 mmol) and DCM (2 mL)followed by p-toluenesulfonyl chloride (0.056 g, 0.294 mmol). Thereaction began to stir at rt for 20 h and was then diluted withsaturated NaHCO₃ and extracted with EtOAc. The EtOAc was then dried withMgSO₄, filtered and concentrated then purified by reverse-phase MPLC(10-100% MeCN:water) to provide pure 3-(tert-butyl)phenethyl4-methylbenzenesulfonate (0.060 g, 0.180 mmol, 92% yield). ¹H NMR (400MHz, CDCl₃): δ 7.70 (d, J=8.4 Hz, 2H), 7.30-7.24 (m, 3H), 7.19 (t,J=7.52 Hz, 1H), 7.14-7.12 (m, 1H), 4.22 (t, J=7.2 Hz, 2H), 2.96 (t,J=7.2 Hz, 2H), 2.43 (s, 3H), 1.29 (s, 9H).

KSC-352-097 (1-(3-(Tert-butyl)phenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the 3-(tert-butyl)phenethyl 4-methylbenzenesulfonate(KSC-352-093) (0.060 g, 0.180 mmol), diphenyl(piperidin-4-yl)methanol(0.048 g, 0.180 mmol) and acetonitrile (1 mL). The TEA (0.038 mL, 0.271mmol) was added and the reaction stirred at 85° C. for 18 h and wascooled to rt then diluted with water and extracted with EtOAc. The EtOAclayer was concentrated and the crude product was purified byreverse-phase MPLC (10-100% MeCN:water) to produce the pure(1-(3-(tert-butyl)phenethyl)piperidin-4-yl)diphenylmethanol (0.065 g,0.152 mmol, 84% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.51-7.47 (m, 4H),7.32-7.27 (m, 4H), 7.23-7.16 (m, 5H), 7.01-6.98 (m, 1H), 3.09-3.04 (m,2H), 2.81-2.76 (m, 2H), 2.61-2.55 (m, 2H), 2.51-2.43 (m, 1H), 2.28 (brs, 1H), 2.10-2.02 (m, 2H), 1.30 (s, 9H), 1.32-1.26 (m, 4H). ¹³C NMR (125MHz, CDCl₃): δ 151.2, 145.9, 140.0, 128.1, 128.0, 126.5, 125.8, 125.71,125.68, 123.0, 79.5, 54.1, 44.2, 34.6, 34.0, 31.6, 31.4, 26.4, 22.6.LCMS Retention time: 4.214 min. LCMS purity 96.4%. HRMS (ESI): m/z calcdfor C₃₀H₃₇NO [M+H]⁺ 428.2875. found 428.2948.

KSC-352-099 3-((Phenylsulfonyl)methylene)oxetane

To an oven-dried vial was added (methylsulfonyl)benzene (0.570 g, 3.65mmol) and the vial was evacuated with argon 3 times. The dry THF (17 mL)was added and the reaction was cooled to 0° C. The 2.5 M BuLi in hexanes(3.21 mL, 8.03 mmol) was added dropwise and the reaction began to stirat 0° C. and stirred for 45 minutes. The diethyl chlorophosphate (0.528mL, 3.65 mmol) was then added at 0° C. and the reaction stirred for 30minutes. The reaction was then cooled to −78° C. and the oxetan-3-one(0.330 mL, 5.15 mmol) was then added dropwise and the reaction stirredfor 2 h. The reaction was then warmed to rt and filtered through asilica plug. The reaction was then concentrated onto silica and purifiedby MPLC (20 min, 0-40% EtOAc:hex) to provide pure3-((phenylsulfonyl)methylene)oxetane (0.579 g, 2.75 mmol, 75% yield). ¹HNMR (400 MHz, CDCl₃): δ 7.91-7.87 (m, 2H), 7.69-7.64 (m, 1H), 7.60-7.55(m, 2H), 6.12 (quintet, J=2.3 Hz, 1H), 5.66-5.63 (m, 2H), 5.30-5.27 (m,2H).

KSC-367-003 2-(4-(3-((Phenylsulfonyl)methyl)oxetan-3-yl)phenyl)ethanol

To a vial was added the chloro(1,5-cyclooctadiene)rhodium(I), dimer(0.012 g, 0.025 mmol) and 1,4-dioxane (10 mL). The 1.5 M aqueous KOH(0.496 mL, 0.745 mmol) was then added and the reaction stirred for 1minute at rt. Then the (4-(2-hydroxyethyl)phenyl)boronic acid (0.103 g,0.621 mmol) and 3-((phenylsulfonyl)methylene)oxetane (KSC-352-099)(0.052 g, 0.248 mmol) in 1 mL dioxane was added and the reaction stirredfor 30 minutes at 100° C. in μwaves. The reaction was then cooled to rtand diluted with EtOAc and washed with 1.0 M HCl. The water layer wasextracted with EtOAc (3×15 mL). The EtOAc layer was collected and thewater was extracted with EtOAc again. The EtOAc layers were combined anddried with MgSO₄, filtered and concentrated then purified byreverse-phase MPLC (30 min, 10-100% MeCN:water) to produce pure2-(4-(3-((phenylsulfonyl)methyl) oxetan-3-yl)phenyl)ethanol (0.063 g,0.190 mmol, 76% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.55-7.52 (m, 2H),7.50-7.46 (m, 1H), 7.36-7.31 (m, 2H), 7.09 (d, J=8.4 Hz, 2H), 7.01 (d,J=8.3 Hz, 2H), 5.03 (d, J=6.4 Hz, 2H), 4.93 (d, J=6.4 Hz, 2H), 4.03 (s,2H), 3.82 (t, J=7.3 Hz, 2H), 2.80 (t, J=7.3 Hz, 2H).

KSC-367-027 4-(diphenylmethylene)piperidine

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.514 g, 1.922mmol) and TFA (4 mL). The reaction stirred at 75° C. at for 24 h and wasconcentrated in vacuo. The residue was then dissolved in DCM (5 mL) andwashed with water (4 mL). The DCM layer was collected and washed withwater (1×5 mL), dried with MgSO₄, filtered concentrated then purified byreverse-phase MPLC (10-100% MeCN:Hex) to produce pure4-(diphenylmethylene)piperidine (0.296 g, 1.187 mmol, 62% yield). ¹H NMR(400 MHz, CDCl₃): δ 7.34-7.28 (m, 4H), 7.25-7.20 (m, 2H), 7.18-7.14 (m,4H), 2.96-2.92 (m, 4H), 2.37-2.33 (m, 4H), 1.75 (br s, 1H).

KSC-367-0324-(4-Benzylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-one

To a vial was added the 4-benzylpiperidine (0.25 ml, 1.422 mmol),1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-one (0.407 g, 1.707 mmol),acetonitrile and TEA (0.297 mL, 2.133 mmol). The reaction stirred at 85°C. for 17 h. The reaction was cooled to rt and diluted with saturatedNaHCO₃ then extracted with EtOAc. The EtOAc was dried with MgSO₄,filtered and concentrated. The crude material was purified byreverse-phase MPLC (10-100% MeCN:water) to produce4-(4-benzylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-one (0.291 g,0.771 mmol, 54% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.91 (d, J=8.6 Hz,2H), 7.47 (d, J=8.5 Hz, 2H), 7.29-7.24 (m, 2H), 7.20-7.16 (m, 1H),7.14-7.11 (m, 2H), 2.96 (t, J=7.2 Hz, 2H), 2.90-2.84 (m, 2H), 2.50 (d,J=7.0 Hz, 2H), 2.36 (t, J=7.3 Hz, 2H), 1.96-1.82 (m, 4H), 1.63-1.56 (m,2H), 1.53-1.46 (m, 1H), 1.34 (s, 9H), 1.28-1.18 (m, 2H).

KSC-367-033 3-(4-(Hydroxydiphenylmethyl)piperidin-1-yl)propan-1-ol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.502 g, 1.878mmol), 3-bromopropan-1-ol (0.204 mL, 2.253 mmol), acetonitrile and TEA(0.393 mL, 2.82 mmol). The reaction stirred at 85° C. for 3 h. Thereaction was cooled to rt and diluted with saturated NaHCO₃ andextracted with EtOAc. The EtOAc layer was concentrated and purified byreverse-phase MPLC (10-100% MeCN:water) to provide3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propan-1-ol (0.395 g, 1.214mmol, 65% yield) as a white solid. ¹H NMR (400 MHz, CDCl₃): δ 7.48-7.44(m, 4H), 7.31-7.27 (m, 4H), 7.20-7.15 (m, 2H), 5.45 (br s, 1H), 3.75 (t,J=5.2 Hz, 2H), 3.12-3.06 (m, 2H), 2.57 (t, J=5.7 Hz, 2H), 2.47-2.36 (m,1H), 1.98-1.91 (m, 2H), 1.70-1.65 (m, 3H), 1.54-1.42 (m, 4H).

KSC-367-036 3-(4-(Hydroxydiphenylmethyl)piperidin-1-yl)propyl4-methylbenzenesulfonate

Prepared with same method at KSC-352-093 using3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propan-1-ol (KSC-367-033)(0.395 g, 1.214 mmol), 4-methylbenzene-1-sulfonyl chloride (0.347 g,1.821 mmol), DCM (5 mL) and TEA (0.508 mL, 3.64 mmol) to produce pure3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propyl4-methylbenzenesulfonate (0.193 g, 0.402 mmol, 33% yield). ¹H NMR (400MHz, CDCl₃): δ 7.69-7.66 (m, 2H), 7.46-7.41 (m, 4H), 7.24-7.19 (m, 4H),7.15-7.05 (m, 4H), 4.26-4.12 (m, 4H), 3.73-3.65 (m, 2H), 3.27-3.18 (m,2H), 2.68-2.60 (m, 1H), 2.47-2.37 (m, 2H), 2.30 (s, 3H), 1.88-1.76 (m,2H), 1.52-1.43 (m, 2H).

KSC-367-0394-(4-Benzylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-ol

Method B: 4-(4-benzylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-one(KSC-367-032) (0.291 g, 0.771 mmol) and MeOH (5 mL) and sodiumborohydride (0.117 g, 3.08 mmol) to produce pure4-(4-benzylpiperidin-1-yl)-1-(4-(tert-butyl)phenyl)butan-1-ol (0.232 g,0.611 mmol, 79% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.35-7.25 (m, 6H),7.21-7.12 (m, 3H), 4.64-4.60 (m, 1H), 3.13-3.07 (m, 1H), 2.94-2.88 (m,1H), 2.54 (d, J=7.0 Hz, 2H), 2.44-2.34 (m, 2H), 2.03-1.93 (m, 2H),1.90-1.38 (m, 10H), 1.31 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ 149.4,142.9, 140.7, 129.1, 128.2, 125.8, 125.4, 125.0, 73.5, 59.0, 54.6, 52.9,42.9, 40.1, 38.0, 34.4, 31.8, 31.6, 31.4, 24.3. LCMS Retention time:4.330 min. LCMS purity 99.1%. HRMS (ESI): m/z calcd for C₂₆H₃₇NO [M+H]⁺380.2875. found 380.2948.

KSC-367-043(1-(3-((4-(Tert-butyl)phenyl)amino)propyl)piperidin-4-yl)diphenylmethanol

To a vial was added the3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propyl4-methylbenzenesulfonate (KSC-367-036) (0.088 g, 0.183 mmol),4-(tert-butyl)aniline (0.035 mL, 0.220 mmol) and TEA (0.038 mL, 0.275mmol) with acetonitrile (3 mL). The reaction began to stir at 85° C. for18 h and was then cooled to rt and diluted with saturated NaHCO₃ thenextracted with EtOAc. The EtOAc was concentrated and purified byreverse-phase MPLC (10-100% MeCN:water) to produce pure(1-(3-((4-(tert-butyl)phenyl)amino)propyl)piperidin-4-yl)diphenylmethanol (0.038 g, 0.083 mmol, 45% yield).¹H NMR (400 MHz, CDCl₃): δ 7.50-7.46 (m, 4H), 7.33-7.28 (m, 4H),7.21-7.16 (m, 4H), 6.51 (d, J=8.7 Hz, 2H), 3.13 (t, J=6.4 Hz, 2H),3.02-2.96 (m, 2H), 2.49-2.41 (m, 3H), 2.15 (br s, 1H), 2.00-1.93 (m,2H), 1.80-1.73 (m, 2H), 1.57-1.43 (m, 5H), 1.27 (s, 9H). ¹³C NMR (125MHz, CDCl₃): δ 146.4, 145.9, 139.7, 128.2, 126.5, 125.9, 125.8, 112.4,79.6, 57.4, 54.2, 44.2, 43.7, 33.8, 31.5, 26.6, 26.2. LCMS Retentiontime: 4.208 min. LCMS purity 100%. HRMS (ESI): m/z calcd for C₃₀H₃₈N₂O[M+H]⁺ 457.6620. found 457.3194.

KSC-367-044(1-(3-(4-(Tert-butyl)phenoxy)propyl)piperidin-4-yl)diphenylmethanol

To a vial was added the3-(4-(hydroxydiphenylmethyl)piperidin-1-yl)propyl4-methylbenzenesulfonate (KSC-367-036) (0.105 g, 0.219 mmol),4-(tert-butyl)phenol (0.039 g, 0.263 mmol) and TEA (0.046 mL, 0.328mmol) with acetonitrile (3 mL). The reaction stirred at 85° C. for 18 hthen diluted with saturated NaHCO₃ and extracted with EtOAc. The EtOAclayer was concentrated and purified by RP MPLC (10-100% MeCN:water) toproduce pure(1-(3-(4-(tert-butyl)phenoxy)propyl)piperidin-4-yl)diphenylmethanol(0.015 g, 0.033 mmol, 15% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.49-7.46(m, 4H), 7.31-7.26 (m, 6H), 7.19-7.14 (m, 2H), 6.81 (d, J=8.8 Hz, 2H),3.96 (t, J=6.4 Hz, 2H), 3.01-2.94 (m, 2H), 2.51-2.40 (m, 3H), 1.99-1.89(m, 5H), 1.52-1.45 (m, 4H), 1.28 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ156.7, 145.9, 143.2, 128.1, 126.5, 126.1, 125.7, 116.3, 113.9, 79.5,66.3, 60.4, 55.4, 54.1, 44.1, 34.0, 31.5, 29.7, 27.0, 26.4. LCMSRetention time: 4.293 min. LCMS purity 95.2%. HRMS (ESI): m/z calcd forC₃₀H₃₇NO₂ [M+H]⁺ 458.6470. found 458.3068.

KSC-367-047 2-(6-(tert-butyl)pyridin-3-yl)ethanol

Prepared by same method as KSC-352-090 with5-bromo-2-(tert-butyl)pyridine (0.303 g, 1.415 mmol), dry THF (10 mL),2.5 M BuLi in hexanes (0.623 mL, 1.557 mmol) and ethylene oxide (1.415ml, 3.54 mmol) (2.5-3.3M solution in THF) to produce pure2-(6-(tert-butyl)pyridin-3-yl)ethanol (0.211 g, 1.177 mmol, 83%yield).). ¹H NMR (400 MHz, CDCl₃): δ 8.40 (br d, J=2.4 Hz, 1H), 7.48(dd, J=8.2 Hz, 2.4 Hz, 1H), 7.26 (dd, J=8.2 Hz, 0.8 Hz, 1H), 3.85 (br t,J=6.0 Hz, 2H), 2.82 (t, J=6.5 Hz, 2H), 2.29 (br s, 1H), 1.33 (s, 9H).

KSC-367-049 2-(6-(tert-butyl)pyridin-3-yl)ethyl 4-methylbenzenesulfonate

Prepared by same method at KSC-352-093 with2-(6-(tert-butyl)pyridin-3-yl)ethanol (KSC-367-047) (0.211 g, 1.177mmol), TEA (0.492 mL, 3.53 mmol) and DCM (2 mL) followed byp-toluenesulfonyl chloride (0.337 g, 1.766 mmol) to provide pure2-(6-(tert-butyl)pyridin-3-yl)ethyl 4-methylbenzenesulfonate (0.260 g,0.780 mmol, 66% yield). ¹H NMR (400 MHz, CDCl₃): δ 8.29 (br d, J=2.4 Hz,1H), 7.69 (d, J=8.3 Hz, 2H), 7.40 (dd, J=8.2 Hz, 2.4 Hz, 1H), 7.30-7.27(m, 2H), 7.23 (dd, J=8.2 Hz, 0.8 Hz, 1H), 4.19 (t, J=6.9 Hz, 2H), 2.92(t, J=6.8 Hz, 2H), 2.43 (s, 3H), 1.34 (s, 9H).

KSC-367-052(1-(2-(6-(Tert-butyl)pyridin-3-yl)ethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.250 g, 0.936mmol), 2-(6-(tert-butyl)pyridin-3-yl)ethyl 4-methylbenzenesulfonate(KSC-367-049) (0.260 g, 0.780 mmol) and acetonitrile (5 mL). The TEA(0.163 mL, 1.170 mmol) was then added and the reaction stirred at 50° C.for 20 h and was then cooled to rt and diluted with saturated NaHCO₃then extracted with DCM. The DCM layer was concentrated and purified byreverse-phase MPLC (10-100% MeCN:water) to produce pure(1-(2-(6-(tert-butyl)pyridin-3-yl)ethyl)piperidin-4-yl)diphenylmethanol(0.308 g, 0.719 mmol, 92% yield). ¹H NMR (400 MHz, CDCl₃): δ 8.39-8.38(m, 1H), 7.51-7.48 (m, 4H), 7.42 (dd, J=8.1 Hz, 2.4 Hz, 1H), 7.32-7.27(m, 4H), 7.25-7.22 (m, 1H), 7.20-7.15 (m, 2H), 3.07-3.01 (m, 2H),2.76-2.71 (m, 2H), 2.57-2.52 (m, 2H), 2.50-2.42 (m, 1H), 2.31 (br s,1H), 2.09-2.01 (m, 2H), 1.57-1.48 (m, 4H), 1.35 (s, 9H). ¹³C NMR (125MHz, CDCl₃): δ 166.9, 148.7, 145.9, 136.3, 132.3, 128.1, 126.5, 125.8,118.6, 79.4, 60.2, 54.0, 44.1, 37.0, 30.3, 30.2, 26.4. LCMS Retentiontime: 3.802 min. LCMS purity 99.5%. HRMS (ESI): m/z calcd for C₂₉H₃₆N₂O[M+H]⁺ 429.2828. found 429.2900.

KSC-367-0531-(4-(Tert-butyl)phenyl)-4-(4-(diphenylmethylene)piperidin-1-yl)butan-1-one

To a vial was added the 4-(diphenylmethylene)piperidin-1-ium2,2,2-trifluoroacetate (KSC-367-027) (0.0475 g, 0.131 mmol),1-(4-(tert-butyl)phenyl)-4-chlorobutan-1-one (0.037 g, 0.157 mmol) andTEA (0.046 mL, 0.327 mmol) in acetonitrile. The reaction stirred at 75°C. for 20 h. The reaction was removed from heat and diluted withsaturated NaHCO₃, extracted with EtOAc. The EtOAc layer was concentratedand purified by reverse-phase MPLC (10-100% MeCN:water) to produce pure1-(4-(tert-butyl)phenyl)-4-(4-(diphenylmethylene)piperidin-1-yl)butan-1-one(0.019 g, 0.042 mmol, 32% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.91 (d,J=8.2 Hz, 2H), 7.46 (d, J=8.3 Hz, 2H), 7.30-7.25 (m, 4H), 7.21-7.17 (m,2H), 7.13-7.10 (m, 4H), 2.99 (t, J=7.2 Hz, 2H), 2.51-2.32 (m, 10H),1.98-1.91 (m, 2H), 1.34 (s, 9H).

KSC-367-055 (1-(4-Chlorophenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the diphenyl(piperidin-4-yl)methanol (0.221 g, 0.827mmol), 1-(2-bromoethyl)-4-chlorobenzene (0.165 g, 0.752 mmol),acetonitrile (4 mL) and TEA (0.157 mL, 1.128 mmol). The reaction stirredat 75° C. for 18 h then was cooled to rt and diluted with saturatedNaHCO₃ and extracted with EtOAc. The EtOAc layer was concentrated andpurified by reverse-phase MPLC (10-100% MeCN:water) to produce pure(1-(4-chlorophenethyl)piperidin-4-yl)diphenylmethanol (0.224 g, 0.552mmol, 73% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.39-7.36 (m, 4H),7.20-7.15 (m, 4H), 7.12-7.09 (m, 2H), 7.08-7.03 (m, 2H), 6.99-6.96 (m,2H), 2.92-2.87 (m, 2H), 2.64-2.58 (m, 2H), 2.43-2.38 (m, 2H), 2.37-2.30(m, 1H), 2.22 (br s, 1H), 1.95-1.88 (m, 2H), 1.44-1.35 (m, 4H). ¹³C NMR(125 MHz, CDCl₃): δ 145.9, 145.8, 138.9, 131.6, 129.9, 128.3, 128.0,126.4, 125.7, 79.4, 79.3, 60.4, 57.0, 44.1, 33.0, 26.4. LCMS Retentiontime: 3.903 min. LCMS purity 99.6%. HRMS (ESI): m/z calcd for C₂₆H₂₈ClNO[M+H]⁺ 406.1859. found 406.1932.

KSC-367-0581-(4-(Tert-butyl)phenyl)-4-(4-(diphenylmethylene)piperidin-1-yl)butan-1-ol

Method B:1-(4-(tert-butyl)phenyl)-4-(4-(diphenylmethylene)piperidin-1-yl)butan-1-one(KSC-367-053) (0.019 g, 0.042 mmol) and sodium borohydride (6.37 mg,0.168 mmol) to produce pure1-(4-(tert-butyl)phenyl)-4-(4-(diphenylmethylene)piperidin-1-yl)butan-1-ol(0.010 g, 0.022 mmol, 52% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.35-7.26(m, 8H), 7.23-7.18 (m, 2H), 7.13-7.10 (m, 4H), 4.68-4.65 (m, 1H),2.66-2.60 (m, 2H), 2.55-2.42 (m, 8H), 2.00-1.94 (m, 1H), 1.90-1.81 (m,1H), 1.73-1.63 (m, 3H), 1.30 (s, 9H). ¹³C NMR (125 MHz, CDCl₃): δ 149.5,142.7, 142.3, 136.4, 134.6, 129.7, 128.0, 126.4, 125.3, 125.0, 73.3,58.7, 55.0, 39.6, 34.4, 31.6, 31.4, 31.1, 24.0, 22.6. LCMS Retentiontime: 4.454 min. LCMS purity 97.1%. HRMS (ESI): m/z calcd for C₃₂H₃₉NO[M+H]⁺ 454.3032. found 454.3104.

KSC-367-066 4-(3-((phenylsulfonyl)methyl)oxetan-3-yl)phenethyl4-methylbenzenesulfonate

Prepared by the same method as KSC-352-093 with2-(4-(3-((phenylsulfonyl)methyl)oxetan-3-yl)phenyl)ethanol (KSC-367-003)(0.104 g, 0.313 mmol), TEA (0.131 mL, 0.939 mmol) and DCM (2 mL)followed by p-toluenesulfonyl chloride (0.089 g, 0.469 mmol) to produce4-(3-((phenylsulfonyl)methyl)oxetan-3-yl)phenethyl4-methylbenzenesulfonate (0.041 g, 0.084 mmol, 27% yield). ¹H NMR (400MHz, CDCl₃): δ 7.75 (d, J=8.3 Hz, 2H), 7.57-7.46 (m, 4H), 7.36-7.32 (m,4H), 7.02-6.96 (m, 3H), 5.00 (d, J=6.4 Hz, 2H), 4.93 (d, J=6.4 Hz, 2H),4.17 (t, J=7.0 Hz, 2H), 4.01 (s, 2H), 2.90 (t, J=7.0 Hz, 2H), 2.45 (s,3H).

KSC-367-069Diphenyl(1-(4-(3-((phenylsulfonyl)methyl)oxetan-3-yl)phenethyl)piperidin-4-yl)methanol

To a vial was added the4-(3-((phenylsulfonyl)methyl)oxetan-3-yl)phenethyl4-methylbenzenesulfonate (KSC-367-066) (0.041 g, 0.084 mmol) anddiphenyl(piperidin-4-yl)methanol (0.025 g, 0.093 mmol) in acetonitrile.The TEA (0.018 ml, 0.126 mmol) was then added and the reaction stirredat 50° C. for 18 h. The reaction was removed from heat and cooled to rtthen diluted with saturated NaHCO₃. The reaction was then extracted withEtOAc (3×5 mL) and the EtOAc layer was dried with MgSO₄, filtered andconcentrated. The crude residue was purified by reverse-phase MPLC (15min, 10-100% MeCN:water) to produce diphenyl(1-(4-(3((phenylsulfonyl)methyl)oxetan-3-yl)phenethyl)piperidin-4-yl)methanol(0.024 g, 0.041 mmol, 49% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.53-7.43(m, 6H), 7.33-7.27 (m, 7H), 7.21-7.16 (m, 2H), 7.04 (d, J=8.2 Hz, 2H),6.96 (d, J=8.2 Hz, 2H), 5.02 (d, J=6.4 Hz, 2H), 4.92 (d, J=6.4 Hz, 2H),4.01 (s, 2H), 3.08-3.02 (m, 2H), 2.74-2.69 (m, 2H), 2.56-2.44 (m, 3H),2.10-2.02 (m, 2H), 1.60-1.50 (m, 5H).

KSC-367-072(1-(4-(3-Methyloxetan-3-yl)phenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added thediphenyl(1-(4-(3-((phenylsulfonyl)methyl)oxetan-3-yl)phenethyl)piperidin-4-yl)methanol(KSC-367-069) (0.024 g, 0.041 mmol) and MeOH (10 mL) and the reaction asheated to 50° C. The magnesium was added in 3 additions (0.016 g, 0.066mmol, 16 equiv) 1.5 h apart. The reaction was removed from heat 1.5 hafter the final magnesium addition, cooled to rt and poured into 1.0 MHCl with ice. The aqueous layer was then extracted with DCM (3×10 mL)and the organic layers were combined and dried with MgSO₄, filtered andconcentrated then purified by reverse-phase MPLC (15 min, 10-100%MeCN:water). To produce(1-(4-(3-methyloxetan-3-yl)phenethyl)piperidin-4-yl)diphenylmethanol(0.0045 g, 10.19 μmol, 24.70% yield) as a light brown solid. ¹H NMR (400MHz, CDCl₃): δ 7.51-7.47 (m, 4H), 7.32-7.27 (m, 4H), 7.21-7.16 (m, 4H),7.13-7.10 (m, 2H), 4.95 (d, J=5.5 Hz, 2H), 4.61 (d, J=5.5 Hz, 2H),3.09-3.02 (m, 2H), 2.81-2.75 (m, 2H), 2.59-2.54 (m, 2H), 2.50-2.42 (m,1H), 2.16 (br s, 1H), 2.09-2.00 (m, 2H), 1.71 (s, 3H), 1.56-1.48 (m,4H). ¹³C NMR (125 MHz, CDCl₃): δ 145.9, 144.1, 138.4, 128.8, 128.2,126.5, 125.8, 125.1, 83.8, 79.5, 60.7, 54.1, 44.1, 43.1, 33.3, 27.8,26.4. LCMS Retention time: 3.652 min. LCMS purity 97.9%. HRMS (ESI): m/zcalcd for C₃₀H₃₅NO₂ [M+H]⁺ 442.2668. found 442.2741.

KSC-367-088 2-(Tert-butyl)phenyl trifluoromethanesulfonate

To a vial was added the 2-(tert-butyl)phenol (1.0 ml, 6.51 mmol) and DCM(4 mL) followed by pyridine (1.053 mL, 13.02 mmol). The reaction wasthen cooled to 0° C. and the triflic anhydride (1.320 mL, 7.81 mmol) wasadded dropwise and the reaction stirred for 2 h. The reaction was thenallowed to warm to rt and was diluted with DCM and quenched with 1.0 MHCl. The DCM layer was collected and washed with saturated NaHCO₃ andbrine. The organic layer was then dried (MgSO₄), filtered and adsorbedto silica and purified by MPLC (0-25% EtOAc:hex) to produce pure2-(tert-butyl)phenyl trifluoromethanesulfonate (1.66 g, 5.88 mmol, 90%yield). ¹H NMR (400 MHz, CDCl₃): δ 7.49-7.46 (m, 1H), 7.37-7.34 (m, 1H),7.31-7.27 (m, 2H), 1.43 (s, 9H).

KSC-381-0092-(2-(Tert-butyl)phenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane

To a flame-dried vial was added molecular sieves and2-(tert-butyl)phenyl trifluoromethanesulfonate (KSC-367-088) (0.475 g,1.683 mmol) and1,1′-Bis(diphenylphosphino)ferrocenedichloropalladium(II) (0.042 g,0.050 mmol). The vial was evacuated with argon 3 times and then dioxane(1 mL), TEA (0.704 mL, 5.05 mmol) and4,4,5,5-tetramethyl-1,3,2-dioxaborolane (0.732 mL, 5.05 mmol) were addedvia syringe. The reaction stirred at reflux (100° C.) for 2 h. Thereaction was then removed from heat and diluted with water thenextracted with DCM (3×5 mL). The DCM layer was then washed again withwater (3×10 mL). The DCM layer was dried with MgSO₄, filtered andconcentrated. The residue was diluted with hexanes and filtered throughMgSO₄ to remove the residual Pd complex. The hexanes layer was thenconcentrated to produce pure2-(2-(tert-butyl)phenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (0.093g, 0.357 mmol, 86% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.45 (dd, J=7.2Hz, 1.2 Hz, 1H), 7.41-7.39 (m, 1H), 7.29 (td, J=7.6 Hz, 2.0 Hz, 1H),7.14 (td, J=7.6 Hz, 2.0 Hz, 1H), 1.41 (s, 9H), 1.38 (s, 12H).

KSC-381-011 1-Bromo-2-(tert-butyl)benzene

To a vial was added the2-(2-(tert-butyl)phenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane(KSC-381-009) (0.180 g, 0.692 mmol) and MeOH (1.5 mL). The copper (II)bromide (0.464 g, 2.075 mmol) was then dissolved in water and added tothe reaction then stirred at 80° C. for 24 h. The reaction was thenremoved from heat and diluted with water and extracted with EtOAc. TheEtOAc was dried with MgSO₄, filtered and concentrated to produce1-bromo-2-(tert-butyl)benzene (0.103 g, 0.483 mmol, 70% yield) as abrown liquid. ¹H NMR (400 MHz, CDCl₃): δ 7.59 (dd, J=8.0 Hz, 1.6 Hz,1H), 7.44 (dd, J=8.0 Hz, 1.6 Hz, 1H), 7.24 (td, J=7.6 Hz, 2.0 Hz, 1H),7.02 (td, J=7.6 Hz, 2.0 Hz, 1H), 1.51 (s, 9H).

KSC-381-015 2-(2-(Tert-butyl)phenyl)ethanol

Prepared by same method as KSC-352-090 with1-bromo-2-(tert-butyl)benzene (KSC-381-011) (0.147 g, 0.690 mmol), dryTHF, 2.5 M BuLi in hexanes (0.303 ml, 0.759 mmol) and then ethyleneoxide (0.690 ml, 1.724 mmol) (2.5-3.3M solution in THF) to produce pure2-(2-(tert-butyl)phenyl)ethanol (0.018 g, 0.101 mmol, 14.64% yield) as aclear oil. ¹H NMR (400 MHz, CDCl₃): δ 7.43-7.37 (m, 1H), 7.23-7.19 (m,1H), 7.18-7.14 (m, 2H), 3.90 (t, J=7.6 Hz, 2H), 3.19 (t, J=7.6 Hz, 2H),1.44 (s, 9H).

KSC-381-018 2-(Tert-butyl)phenethyl 4-methylbenzenesulfonate

Prepared according to same procedure as KSC-352-093 with2-(2-(tert-butyl)phenyl)ethanol (KSC-381-015) (0.018 g, 0.101 mmol), TEA(0.042 mL, 0.303 mmol) and DCM (2 mL) followed by p-toluenesulfonylchloride (0.029 g, 0.151 mmol) to provide pure 2-(tert-butyl)phenethyl4-methylbenzenesulfonate (0.022 g, 0.066 mmol, 66% yield) and a clearoil. ¹H NMR (400 MHz, CDCl₃): δ 7.76 (d, J=8.0 Hz, 2H), 7.37-7.31 (m,3H), 7.17-7.04 (m, 3H), 4.19 (t, J=7.6 Hz, 2H), 3.25 (t, J=7.6 Hz, 2H),2.44 (s, 3H), 1.33 (s, 9H).

KSC-381-020 (1-(2-(Tert-butyl)phenethyl)piperidin-4-yl)diphenylmethanol

To a vial was added the 2-(tert-butyl)phenethyl 4-methylbenzenesulfonate(KSC-381-018) (0.022 g, 0.066 mmol), diphenyl(piperidin-4-yl)methanol(0.018 g, 0.066 mmol) and acetonitrile. The TEA (0.014 mL, 0.099 mmol)was added and the reaction stirred at 85° C. for 20 h. The reaction wascooled to rt and diluted with water then extracted with EtOAc. The EtOAclayer was concentrated and the crude product was purified byreverse-phase MPLC (10-100% MeCN:water) to produce the pure(1-(2-(tert-butyl)phenethyl) piperidin-4-yl)diphenylmethanol (0.013 g,0.030 mmol, 46% yield). ¹H NMR (400 MHz, CDCl₃): δ 7.51-7.48 (m, 4H),7.37-7.28 (m, 5H), 7.21-7.09 (m, 5H), 3.15-3.04 (m, 4H), 2.65-2.60 (m,2H), 2.52-2.42 (m, 1H), 2.17-2.08 (m, 3H), 1.60-1.53 (m, 4H), 1.41 (s.9H). ¹³C NMR (125 MHz, CDCl₃): δ 147.7, 145.9, 138.5, 132.1, 128.2,126.5, 126.1, 125.9, 125.84, 125.79, 79.5, 61.7, 54.2, 44.2, 35.7, 31.7,29.7, 26.4. LCMS Retention time: 4.200 min. LCMS purity 99.3%. HRMS(ESI): m/z calcd for C₃₀H₃₇NO [M+H]⁺ 428.2875. found 428.2948.

Bacterial Strains and Conditions

All strains used to evaluate the antimicrobial activity of terfenadineand corresponding structural derivatives are shown below in Table 5. S.aureus strain UAMS-1 is an osteomyelitis clinical isolate (Gillaspy etal. “Role of the accessory gene regulator (agr) in pathogenesis ofstaphylococcal osteomyelitis. Infect. Immun. 63:3373-3380 (1995)),whereas ciprofloxacin-resistant strains CRC118 and CRC61 are spontaneousciprofloxacin-resistant derivatives of UAMS-1 that were selected bygrowth on Mueller-Hinton agar (MHA) (Becton, Dickinson & Company,Franklin Lakes, N.J.) at 1.5×MIC ciprofloxacin (0.75 μg/ml).

TABLE 5 Bacterial Strains Used in Terfenadine Study Species StrainSource S. aureus UAMS-1 1 CRC61 Dunman Lab, URMC CRC118 Dunman Lab, URMC

Minimum Inhibitory Concentration (MIC) Testing

Minimum Inhibitory Concentration (MIC) testing was performed todetermine the minimum concentration of test compound that is necessaryto inhibit visible growth of bacteria according to Clinical andLaboratory Standards (CLSI) guidelines (Clinical and LaboratoryStandards Institute (CLSI). Methods for Dilution AntimicrobialSusceptibility Tests for Bacteria That Grow Aerobically; ApprovedStandard—Ninth Edition. CLSI document M07-A9. 2012. Clinical andLaboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne,Pa. 19087-1898 USA.) To do so, 10⁵ colony forming units of an overnightbacterial culture were seeded into individual wells of 96-wellround-bottom microtiter plates containing 88 μl of MHB medium. To thefirst column, 2 μl of the test compound's corresponding solvent was alsoadded to each well (negative control). To the next ten columms, 2 μl ofthe test compound (dissolved in DMSO for terfenadine and its derivativesor sterile water for ciprofloxacin) was added in increasing 2-foldincrements of 0.5 μg/ml to 256 μg/ml to each successive well. Each testcompound was evaluated in duplicate. Plates were incubated at 37° C.incubator for 16 h at which point the minimum inhibitory concentrationwas determined to be the lowest concentration of test compound thatinhibited bacterial growth, as judged by the unaided eye.

S. aureus DNA Gyrase Supercoiling Assay (TopoGEN)

The gyrase supercoiling assays were performed to determine if testcompounds interfered with S. aureus DNA gyrase activity, following themanufacturer's recommendations (TopoGEN). Reactions (20 μl) containedkit provided assay buffer, ATP, potassium glutamate, relaxed plasmid DNA(0.4 μg/ml), 2 Units of S. aureus DNA gyrase, and various amounts oftest compound. Reactions were incubated at 37° C. for 30 min and thenstopped by the addition of 10% SDS, filtered through 0.025 μm Milliporemembrane filters in a 10 mM Tris-HCl buffer (pH 8), and electrophoresedin a 1% agarose TAE gel. Gels were stained with 0.5 μg/ml ethidiumbromide and images were analyzed using densitometry (Image J, NIH). TheIC₅₀ values for each test compound was determined to be the compoundconcentration that inhibited S. aureus DNA gyrase activity by 50%.

S. aureus Topoisomerase IV Decatenation Assay (Inspiralis)

A Topoisomerase IV assay was performed on test compounds to determine ifthey interfered with the ability of S. aureus topoisomerase IV todecatenate kDNA, according to the recommendations of the manufacturer(Inspiralis). To do so, 0.25 U S. aureus topoisomerase IV enzyme wasmixed with 200 ng kDNA in kit provided reaction buffer, in the absenceor presence of various concentrations of test compounds at 37° C. for 30min. The reaction was stopped by the addition of STEB stop buffer and 30μl of 24:1 chlorform:isoamyl alcohol (total volume 90 μl). Reactionproducts then electrophoresed in a 1% agarose TAE gel, stained with 0.5μg/ml ethidium bromide and images were analyzed using densitometry(Image J, NIH). The IC₅₀ value for each test compound was determined asthe compound concentration that inhibited S. aureus topoisomerase IVactivity by 50%. The results are set forth in Table 6.

TABLE 6 ¶    

Mole- Topo- Com- KUC- cular- MIC- Gyrase- isomeras- pound- Registry-Weight- (μg/ IC50 iv-IC50- #o Numbero (g/mol)o Structureo mL)o (μM)o(μM)o Terfena- dineo N/Ao 471.673

 16o  190.00o  206.67o  1o KSC- 335- 012o 457.647

 16o  126.67o  273.33o  2o KSC- 335- 013o 485.7o

 16o  100.00o  133.33o  3o KSC- 335- 014o 471.673¶

  8o  330.00o  103.33o  4o KSC- 335-015o 450.12o

 64o  133.33o >333o  5o KSC- 335- 016o 457.647o

  8o  133.33o  333.88o  6o KSC- 335- 021o 415.567o

 256o >500o >333o  7o KSC- 335-030o 445.593o

 256o >500o >333o  8o KSC- 335- 031o 433.558o

 128o >500o >333o  9o KSC- 335- 032o 494.463o

  32o >500o >333o 10o KSC- 335- 041o 455.674o

  8o 93.33o 320.00o 11o KSC- 335- 007o 469.658o

  8o  93.33o  110.00o 12o KSC- 335- 069o 471.673o

 16o  126.67o  100.00o 13o KSC- 335- 070o 471.673o

 16o  410.00o  110.00o 14o KSC- 335- 077o 429.594o

 128o >500o >333o 15o KSC- 335- 080o 515.683o

 128o >500o >333o 16o KSC- 335- 081o 501.656o

>256o >500o >333o 17o KSC- 342- 010o 443.62o

 16o  73.33o  100.00o 18c KSC- 342- 017o

  ¶ 473.603

 256o  440.00o >333o 19 KSC- 342- 021 459.577

>256 >500 >333 20 KSC- 342- 006 441.604

>256  73.33  246.67 21 KSC- 335- 008 455.631

 32  90.00  133.33 22 KSC- 342- 080 441.647

  8  93.33  100.00 23 KSC- 342- 081 427.621

  8  93.33  146.67 24 KSC- 342- 088 470.689

 16 >500 >333 25 KSC- 348- 002 491.663

  8  13.33  100.00 26 KSC- 348- 050 413.595

  8  90.00  213.33 27 KSC- 348- 049 427.578

>256  93.33 >333 28 KSC- 348- 058 401.541

 256 >333 >333 29 KSC- 352- 060 416.512

 64  263.33 >333 30 KSC- 352- 061 450.411

 16  246.67  260.00 31 KSC- 352- 066 395.577

 128 >333 >333 32 KSC- 352- 063 447.611

  8  80.00  133.33 33 KSC- 352- 064 448.599

 32  93.33  226.67 34 KSC- 352- 069 386.529

 256  93.33 >333 35 KSC- 352- 065 448.599

 32  250.00  280.00 36 KSC- 352- 075 414.582

 256 >333 >333 37 KSC- 352- 082 439.513

 16  16.67 >333 38 KSC- 352- 088 389.505

 128 >333 >333 39 KSC- 352- 097 427.621

 16  93.33 >333 40 KSC- 367- 039 379.578

 64 >333 >333 41 KSC- 367- 043 456.662

  8  193.33  250.00 42 KSC- 367- 044 457.647

  8  100.00 >333 43 KSC- 367- 052 428.609

 64 >333 >333 44 KSC- 367- 055 405.96

 32  260.00  266.67 45 KSC- 367- 072 441.604

>256 >333 >333

indicates data missing or illegible when filed

1. A method of treating or preventing an infection in a subject with orat risk of developing an infection, the method comprising administeringto the subject a a compound of Formula I

wherein R represents hydrogen or hydroxyl and R¹ represents hydrogen, orR and R¹ taken together form a second bond between the carbon atomsbearing R and R¹; R² represents hydrogen or phenyl; n is zero or apositive whole integer of from 1 to 4; X represents CH₂, CHOH, NH, C═O,CHNR³R⁴, where R³ and R⁴ independently are hydrogen or lower alkyl; andZ represents thienyl, pyridinyl, substituted pyridinyl, phenyl orsubstituted phenyl wherein the substituents on the substituted pyridinylor substituted phenyl are selected from phenyl, pyridinyl, nitro, ahalogen atom, such as chlorine, fluorine, bromine, or iodine, a straightor branched lower alkyl chain of from 1 to 4 carbon atoms, a loweralkoxy group of from 1 to 4 carbon atoms, amino, a mono ordi(lower)alkylamino group, a saturated monocyclic heterocyclic groupsuch as pyrrolidino, piperidino, morpholino, orN-(lower)alkylpiperazino, or a group having the structure —COOR⁵,—CR⁶R⁷COOR⁵, —CF₃, CHF₂, CH₂F, or

where R⁵ is hydrogen or lower alkyl, and R⁶ and R⁷ independently arehydrogen or methyl or a pharmaceutically acceptable salt thereof. 2-4.(canceled)
 5. The method of claim 1, wherein the infection is abacterial infection, a viral infection, a parasitic infection or afungal infection.
 6. The method of claim 1, wherein the infection is arespiratory infection.
 7. The method of claim 1, wherein the infectionis a gastrointestinal infection.
 8. The method of claim 1, wherein theinfection is a skin infection.
 9. The method of claim 5 wherein thebacterial infection is selected from the group consisting ofEnterobacterium faecium, Staphylococcus aureus, Klebsiella pneumonia,Acinebacter baumannii, Pseudomonas aeruginosa and Enterobacter sp. 10.The method of claim 5, wherein the bacterial infection is a small colonyvariant bacterial infection.
 11. A method of treating or preventing abacterial infection in a subject with or at risk of developing abacterial infection, the method comprising administering terfenadine tothe subject.
 12. (canceled)
 13. A method of removing or preventingbiofilm formation on a surface, the method comprising administering to abiofilm containing surface or a surface susceptible to biofilm formationan effective amount of a compound of Formula I

wherein R represents hydrogen or hydroxyl and R¹ represents hydrogen, orR and R¹ taken together form a second bond between the carbon atomsbearing R and R¹; R² represents hydrogen or phenyl; n is zero or apositive whole integer of from 1 to 4; X represents CH₂, CHOH, NH, C═O,CHNR³R⁴, where R³ and R⁴ independently are hydrogen or lower alkyl; andZ represents thienyl, pyridinyl, substituted pyridinyl, phenyl orsubstituted phenyl wherein the substituents on the substituted pyridinylor substituted phenyl are selected from phenyl, pyridinyl, nitro, ahalogen atom, such as chlorine, fluorine, bromine, or iodine, a straightor branched lower alkyl chain of from 1 to 4 carbon atoms, a loweralkoxy group of from 1 to 4 carbon atoms, amino, a mono ordi(lower)alkylamino group, a saturated monocyclic heterocyclic groupsuch as pyrrolidino, piperidino, morpholino, orN-(lower)alkylpiperazino, or a group having the structure —COOR⁵,—CR⁶R⁷COOR⁵, —CF₃, CHF₂, CH₂F, or

where R⁵ is hydrogen or lower alkyl, and R⁶ and R⁷ independently arehydrogen or methyl or a pharmaceutically acceptable salt thereof. 14.The method of claim 13, wherein the biofilm comprises one or more ofStaphylococcus aureus, Pseudomonas aeuroginosa, Staphylococcusepidermidis, Escherichia coli or Acinetobacter baummanii.
 15. A methodof identifying an antimicrobial agent comprising: a) contacting abacterial culture with a test agent; b) and measuring adenylate kinaserelease in the supernatant of the bacterial culture, wherein an increasein adenylate kinase release as compared to a control indicates that thetest compound is an antimicrobial agent.
 16. The method of claim 15,wherein the bacterial culture is selected from the group consisting ofStaphylococcus aureus, Pseudomonas aeuroginosa, Staphylococcusepidermidis, Escherichia coli or Acinetobacter baummanii.
 17. The methodof claim 5, wherein the parasitic infection is not malaria.